2007 Research Award: $86,250

“Mitochondrial calcium signaling and organelle dysfunction in mitochondrial diseases: molecular determinants and regulatory mechanisms.”
An important topic in the study of cell biology has to do with how cells respond to a changing environment in order to keep constant certain factors necessary for their survival. At the same time, the life spans of individual cells are determined by genes that will ultimately cause apoptosis (programmed cell death). Of course, cells can also die prematurely because of disease. Biologists are interested in how these different aspects of a cell’s existence are regulated. A key insight comes from the role that ionic calcium plays as a signal to activate specific processes within the cell. It is also known that build-up of calcium within mitochondria precedes the changes that result in cell death by apoptosis and also premature death due to damage by certain toxins.

Dr. Paolo Pinton and colleagues at the University of Ferrara are conducting UMDF-funded research to investigate calcium balance in mitochondria. This important fundamental research project will increase our understanding of how calcium functions as a mitochondrial regulatory molecule. Because changes in calcium concentration have been linked to certain mitochondrial diseases, it will also help to identify cellular disease mechanisms associated with mitochondrial disorders.

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2007 Research Award: $157,450

“Genotype-Phenotype Correlation and Genetic Modifiers in Barth Syndrome.” Barth syndrome is a rare genetic disorder that causes generalized muscle weakness, affecting both the muscles of the heart and of the musculoskeletal system. The disease also prevents normal function of a specific population of immune system cells and has a high rate of infant mortality. The effects of Barth syndrome on various organs in the human body are due to impaired mitochondria, the energy-providing organelles found in most cells. Muscles and other tissues depend upon the ready availability of mitochondrial ATP in order to function normally. Barth’s patients are thus at a significant disadvantage because they cannot convert adequate amounts of energy from their food into ATP for use by their cells. Because there are only a few patients available for research at any time, it is necessary to use animal models for detailed study of this disease.

 In a UMDF-funded project, Dr. Ren and colleagues at New York University are using fruit flies as a research model for the study of Barth syndrome. Discovery in these insects of genetic modifiers that affect the syndrome’s development will shed light on its pathology and could also point to potential therapies for other mitochondrial disorders.

2007 Research Award: $98,340

“Mitochondrial DNA synthesis and Krebs (tricarboxylic acid) cycle: the succinyl-CoA synthase.”
By now, many people are aware that their DNA contains the genetic information required for the cells in their body to function normally. DNA is actually a code that spells out the structure of a cell’s proteins and most of this information is housed inside the cell’s nucleus. An additional level of complexity was added to the understanding of mitochondrial function when it was discovered that mitochondria also contain DNA. Mitochondria are the primary source of ATP, the high-energy molecule used by cells to carry out a variety of activities. How this organelle is produced in the cell when the genetic instructions for its manufacture are located in two separate cell compartments makes for a complex story. It was recently discovered that mitochondrial disease in some infants is actually due to mitochondrial DNA (mtDNA) depletion, progressive loss of the DNA normally present in their mitochondria. The effects of this depletion are widespread, impairing the function of multiple organs such as the brain, liver, and heart.

Dr. Saada and her colleagues at Hadassah - Hebrew University Medical Center are conducting UMDF-funded research using cells derived from patients with mtDNA depletion. Previous research linked a type of brain mitochondrial disease with mtDNA depletion due to a specific genetic mutation. Her goal is to understand the pathophysiology of this process and to move towards identifying methods for reversing it.

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2007 Research Award: $100,000

“Determination of the nuclear transcriptional responses that affect animal physiopathology upon impaired mitochondrial respiratory chain function.”
For over forty years, biological researchers have used the roundworm Caenorhabditis elegans as a model for understanding the genetic control of development. Its size (a little more than a millimeter in length) and rapid growth (less than four days to reach maturity) makes it an ideal organism for laboratory investigations in the fields of genetics, molecular biology, and neurobiology. Like humans, C. elegans relies upon mitochondria to supply most of its ATP for energy-requiring processes. Also similar to humans, specific mutations in this roundworm’s DNA have been linked with mitochondrial disorders.

Dr. Walter and her colleagues in the Department of Molecular Biology and Genetics at Cornell are conducting UMDF-funded research that is breaking new ground in characterizing how changes in gene expression in C. elegans result in abnormal mitochondrial energy metabolism. As with humans, impaired mitochondrial function can result in a diverse array of functional outcomes and disabilities. It is thus expected that this project will uncover underlying processes that are also at work in human mitochondrial disease.

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2007 Research Award: $94,481

“Increased mitochondrial biogenesis as therapy to mitochondrial myopathies.”
A large percentage of the energy that we derive from the food we eat is used up by our skeletal muscles. These organs have a high metabolic rate that requires a constant supply of ATP, the energy currency that we spend for the many activities necessary for life. Most of this ATP is made available by aerobic (oxygen-requiring) metabolism in mitochondria, which are present in large numbers in the muscles of normal individuals. It is not surprising, then, that patients with various mitochondrial disorders often experience severe muscle weakness and also fatigue easily. What can be done to increase the number of normal mitochondria in patients whose muscles are so starved for energy?

In a UMDF-funded project, Dr. Wenz and colleagues at the University of Miami are determining whether mice with a defective mitochondrial enzyme will improve when they are provided with a gene that increases the number of mitochondria in their muscle cells. These mice, which were developed in her lab, have a progressive disorder leading to greatly impaired muscle function. Investigations with this animal model will hopefully point towards an effective therapy. This is important because of the current scarcity of treatments for mitochondrial disease in humans.

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Brian H. Robinson, PhD
Hospital for Sick Children, Canada

2006 Research Award: $125,000

"High throughput screening for mitochondrial enhancers"
The human brain constantly requires an amount of energy that is far out of proportion to its size. Any factor that decreases energy metabolism in the brain, such as impairment of the mitochondria’s capacity to produce ATP, would have severe consequences both on brain development and function. Drug treatment of neurological disorders is always a challenge because of the blood-brain barrier, the purpose of which is to protect the neurons, brain functional cells, from potential toxins in the blood. The problem with developing effective treatments for brain disorders is that many drugs which might prove effective are denied access into neurons by this barrier. Development of a drug that would stimulate mitochondria to produce ATP at a maximum rate and could also successfully cross into the brain to reach neurons possessing impaired mitochondria is an ongoing challenge.

Dr. Brian Robinson and his colleagues at the Metabolism Research Programme of the Hospital for Sick Children in Toronto are conducting UMDF-funded research to identify chemicals that can cross the blood-brain barrier for their ability to increase ATP production by mitochondria. They are using a cell-based assay allowing rapid screening of a large number of drugs that might enhance the synthesis of mitochondrial respiratory enzymes. This is important because the class of compounds being studied could readily cross into the brain to potentially treat serious neurological diseases in children.

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Thomas W. O’Brien, PhD
University of Florida

2006 Research Award: $125,000

"Mitochondrial ribosomal proteins: candidate genes for mitochondrial disease"
A cell’s DNA is important because of the information it carries. This genetic code contains the directions for manufacturing proteins in the cell. The sequence of events that leads to the formation of these proteins begins with transcribing the DNA into messenger RNA (mRNA). Important organelles called ribosomes direct the subsequent translation of the mRNA code into proteins. If mitochondria are going to provide adequate amounts of energy, then their own ribosomes must function correctly to direct the continued replacement of ATP-producing enzymes. While evidence suggests that incorrectly formed ribosomes may lead to certain mitochondrial diseases, little is known about their components and how these ribosomal proteins contribute to the overall process of translating mRNA into enzymes required by the mitochondria.

Dr. Thomas O’Brien and colleagues at the University of Florida are conducting UMDF-funded research to further our understanding of the functional significance of a number of ribosomal proteins in mitochondria. This research is important because their lab is especially interested in identifying proteins associated with specific mitochondrial diseases.

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Håkan Westerblad, MD, PhD
Karolinska Institute, Sweden

2006 Research Award: $122,720

"Mechanisms of muscle dysfunction studied in mouse models of mitochondrial myopathies"
It is well known that cells require the organelles called mitochondria to function normally so as to maintain a constant supply of energy. When a patient’s mitochondria are defective, organs needing large amounts of energy, such as skeletal muscle, are especially affected. Because both the maintenance of healthy muscles and the regulation of their contraction are so complex, the relationships between mitochondrial disease and muscle pathology are also likely to be complex. Obtaining a detailed picture of these disease mechanisms in humans is very difficult. What is needed is a means of linking specific mitochondrial defects with specific areas of muscle dysfunction in isolated muscles obtained from individuals with known mitochondrial defects.

Dr. Håkan Westerblad and colleagues at the Department of Physiology and Pharmacology of the Karolinska Institute have developed animal models that allow them to answer such questions. Conducting UMDF-funded research, they are comparing various aspects of function in muscle from normal mice and from mice with different types of mitochondrial defects. They are investigating regulation of muscle contraction, force generation by muscles, and factors that can lead to the death of individual muscle cells. The importance of this research lies in the detailed information that will be gained about exactly how abnormal mitochondria cause muscle dysfunction. Once information concerning specific disease mechanisms has been acquired, it should provide insight into promising treatment strategies.

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Haya Lorberboum-Galski, PhD
Hebrew University of Jerusalem

2006 Research Award: $115,000

"Enzymereplacement therapy: A novel approach for treating a mitochondrial disease-LAD deficiency"
The many different medical conditions that are placed under the category “mitochondrial disease” have in common the disruption of one or more components of the energy “machinery” of the mitochondria. This machinery is made up of a number of enzymes that are crucial to the production of the energy molecule ATP. If even one of these enzymes is defective or absent, then ATP production may be greatly reduced, with serious consequences for organs such as the brain and skeletal muscle that depend upon large amounts of ATP for normal development and activity. Lipoamide dehydrogenase (LAD) is an important component of three enzyme complexes in mitochondria and LAD deficiency is an inherited disease that disrupts normal mitochondrial function.

Dr. Lorberboum-Galski and colleagues at the Department of Cellular Biochemistry and Human Genetics at Hebrew University are conducting UMDF-funded research to investigate the possibility that enzyme replacement therapy can be used to treat LAD deficiency. They are developing methods for placing the normally functioning enzyme into cultured cells of mitochondrial disease patients and also into the mitochondria of laboratory mice that are afflicted with the disease. This is a promising approach because there are currently no cures for mitochondrial disease and such research could lead to therapies that would correct its fundamental causes.

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Zaza Khuchua, PhD
Vanderbilt University Medical Center, Tennessee

2006 Research Award: $110,000

"Animal models of human Barth syndrome, a mitochondrial cardiolipin disorder"
Barth syndrome is a rare genetic disorder that causes generalized muscle weakness, affecting both the muscles of the heart and of the musculoskeletal system. The disease prevents normal function of a specific population of immune system cells and has a high rate of infant mortality. The similarity of effects that the disease has on very different organs in the human body can be explained by how it affects mitochondria, energy-providing organelles found in most cells. Previous studies have shown that the muscle mitochondria in Barth patients are deficient in cardiolipin, a compound that is an essential component of the mitochondrial inner membrane. Production of the energy needed for cell activity is impaired in the absence of normal cardiolipin levels.

Dr. Zaza Khuchua and colleagues at Vanderbilt University Medical Center have developed promising animal models of human Barth syndrome. Using research funds provided by the United Mitochondrial Disease foundation, they will study a previously developed fish model of Barth syndrome and will complete development of a mouse model of the same disease. These are significant developments because the use of animal models will allow extensive research on the syndrome’s basic pathology without having to rely upon human subjects.

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Stephane Chiron, PhD
University of California-San Diego

2006 Research Award: $98,500

"Utilization of fission yeast as a model for mitochondrial morphology: a new approach to discover novel genes involved in animal cells"
The cells that make up the human body are packed with organelles, complex machinery responsible for virtually all life processes. These functional components are dynamic and mobile, not the static structures often suggested by pictures in biology textbooks. Mitochondria are essential organelles responsible for maintaining normal energy levels in cells and previous research has shown that specific transport systems are dedicated to moving them within the cell interior. Other research has suggested that abnormal movement and shaping of mitochondria are associated with certain muscle and nervous system diseases in humans. Detailed studies of the internal systems responsible for distributing and positioning mitochondria within cells could shed light on disease mechanisms common to some mitochondrial diseases.

Dr. Stephane Chiron and colleagues at U.C.-San Diego are conducting UMDF-funded research seeking to further understand how mitochondrial movement is regulated. They are using a species of yeast that has the same microtubule transport system for movement of mitochondria as is found in human cells. Such yeast cells are easily cultured and maintained in the lab and, because of their similarities with human cells, will yield data that is applicable to human mitochondrial disease.

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Michael J. Palladino, PhD
University of Pittsburgh

2006 Research Award: $98,457

"Developing therapies for mitochondrial disease"
Mitochondrial diseases are relatively rare and a single clinician may encounter only a small number of such patients over a long period of time. Developing a profile of the “typical” patient is challenging also because any two individuals with the same disorder may follow different courses and experience signs and symptoms that differ in both type and severity. It is therefore difficult to collect a large number of similar mitochondrial disease patients for clinical research in order to assess the effectiveness of promising treatments. Fruit flies have many genetic features in common with humans and have been used for decades to uncover aspects of human disease. Using modern biotechnology to produce fruit flies with specific genetic anomalies that lead to mitochondrial disease would be very helpful in identifying underlying disease mechanisms.

Dr. Michael Palladino and his colleagues at the University of Pittsburgh are using fruit flies with mutated mitochondria to measure the effectiveness of treatments for diseases that cause progressive deterioration of the nervous and muscular systems. The mitochondrial dysfunction in these animals causes movement disorders and decreases their life spans. The importance of this UMDF-funded research is that it will allow wide-spread screening to determine the efficacy of specific drug therapies in genetically similar populations with mitochondrial disease.

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Doron Rapaport, PhD
University of Tuebingen, Germany

2006 Research Award: $98,000

"Defective biogenesis of mitochondrial beta-barrel proteins as a cause for Mohr-Tranebjaerg syndrome"
Mitochondria are complex cell organelles whose manufacture is directed by the DNA code contained in genes of both the nucleus and of the mitochondria themselves. Because the majority of mitochondrial components are encoded for in nuclear DNA, their assembly involves the transport of proteins from the interior of the cell into the mitochondria. Anything that interrupts this complicated sequence of events has the potential to impair mitochondrial function, leading to impaired mitochondrial energy metabolism. The consequences of such impairment can be seen in Mohr-Tranebjaerg syndrome (MTS), a disorder of the nervous system that results in the development of deafness, blindness, mental retardation and dysfunctional movement.

Doron Rapaport and his colleagues at the University of Tuebingen are conducting UMDF-funded research to investigate the link between a mutation associated with MTS and the insertion of nuclear-encoded proteins into the mitochondrial membrane. This research is important because it will provide insights into the normal assembly of mitochondria, as well as into the pathology of a complex mitochondrial disease.

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Vishal Gohil, PhD
Massachusetts General Hospital

2006 Research Award: $88,850

"Molecular signatures of mitochondrial disorders"
It should not be surprising to learn that our muscles use up a large percentage of the energy we derive from our food. Muscle cells have such a high metabolic rate that that they burn a significant number of calories even when we are asleep. It is the job of mitochondria in these muscle cells to make energy available in the form of ATP on a reliable basis. ATP is the currency that is “spent” for energy-requiring activities in the human body and therefore mitochondria are present in large numbers in normal muscle cells. Conversely, when patients have widespread energy dysfunction associated with abnormal mitochondria, the muscles are especially vulnerable.

Dr. Vishal Gohil and colleagues at the MGH Center for Human Genetic Research are conducting UMDF-funded research to investigate the effects of malfunctioning mitochondria on mammalian muscle function. They are developing research models for about a dozen different human mitochondrial diseases by interfering with the expression of DNA in a mouse muscle cell line. This approach is important because it will allow comprehensive laboratory study of a number of mitochondrial diseases and their associated defects. Their findings should provide insight into appropriate treatments for mitochondrial disease in humans.

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John Gordon Lindsay, PhD
University of Glasgow, Scotland

2006 Research Award: $43,494

"Enzymatic, assembly and genetic studies on the human pyruvate dehydrogenase multi-enzyme complex"
The machinery inside living cells is very complex and includes a variety of enzymes that regulate the rates at which chemical reactions occur. The reactions that take place within mitochondria are especially important because they are crucial to maintaining normal levels of the energy molecule ATP. The process whereby the chemical energy in glucose is ultimately transferred to ATP begins in the cytosol outside of the mitochondria and results in the formation of an intermediate molecule called pyruvate. While a relatively small amount of ATP becomes available during the breakdown of a glucose molecule into two pyruvates, obtaining the maximum amount of energy available from glucose requires that the pyruvates enter the mitochondria for further processing. Once inside, the pyruvate molecules undergo a chemical reaction that is catalyzed by pyruvate dehydrogenase complex (PDC), an important mitochondrial enzyme.

Dr. Lindsay and his colleagues at the Institute of Biomedical and Life Sciences of the University of Glasgow are conducting UMDF-funded research using modified bacteria that contain mutated forms of PDC in order to determine how changes in the complex’s function can result in mitochondrial disease. If the enzyme is defective, then numerous other reactions that follow the one catalyzed by PDC will not occur and mitochondrial production of ATP will be impaired. This research is important because PDC mutations are suspected in a large number of metabolic disorders in humans.

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Patrick F. Chinnery, PhD
University of Newcaslte upon Tyne, UK

2005 Research Award: $162,878

"The Population prevalence of ten mtDNA mutations"
Each of us depends upon our own mitochondria to derive energy, in the form of ATP, from the food that we eat. Because ATP has to be readily available for use by our cells the mitochondrial enzyme systems responsible for its production are constantly working. Unfortunately, this complicated machinery can malfunction, sometimes because the mitochondrial enzyme complexes were improperly assembled. What specifically goes wrong with the mitochondria to cause this in patients with mitochondrial disease? We know that the enzyme complexes in the mitochondria are manufactured by following the directions contained in the genetic code in the form of DNA. Most of this DNA is present in the nucleus, but some is actually inside the mitochondria themselves. One possible cause of mitochondrial malfunction is that the mitochondrial DNA (mtDNA) contains a mutated faulty genetic code. But how common are these mutations in the general population?

Professor Patrick F. Chinnery and his team at the University of Newcastle Upon Tyne in the U.K., are conducting UMDF-funded research to determine the prevalence of mutations in mtDNA in the general population. They are comparing these findings with the number of mutations identified in mitochondria from individuals living in the same region who are affected with mitochondrial disease. It is suspected that mtDNA mutations are more common than is currently understood. Dr. Chinnery’s study will lay the groundwork for developing a more accurate assessment on their incidence in the population at large.

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Michael Frohman, MD PhD
Stony Brook University, New York

2005 Research Award: $141,027 

"MitoPLD, a novel enzymatic regulator of mitochondrial morphology and fusion"
Mitochondria are complicated structures that, in some ways, are like miniature cells within a cell. Unlike any other cell organelles, mitochondria contain their own DNA, which is used, along with DNA in the nucleus, to direct the manufacture and maintenance of their own enzyme systems. It is the mitochondrial enzymes that oversee a complex set of reactions which transfer energy from the food we eat to the widely used energy molecule known as ATP. Although they are typically depicted in biology textbooks as kidney-bean shaped structures, mitochondria can merge or fuse with each other to form much longer, complex organelles. Previous studies have shown that conditions which favor mitochondrial fusion are necessary for them to carry out energy metabolism at normal levels. Factors that block fusion of mitochondria lead to fragmentation of the organelles and impair their activity. Fusion apparently helps to keep some defective mitochondria functional by providing the correct DNA genetic code for enzyme synthesis.

Dr. Michael Frohman and his team at Stony Brook University in New York are conducting UMDF-funded research that will add to our understanding of why the fusion of mitochondria is so crucial to their normal operation. They have discovered a new gene, called MitoPLD, which regulates mitochondrial fusion and are determining how the gene works. This is important because defective fusion of mitochondria is linked to specific mitochondrial diseases of the nervous system, and perhaps as well as those of other body systems.

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Elena Rugarli, MD
Telethon Institute of Genetics and Medicine, Italy

2005 Research Award: $126,500 

"The Mechanism of Mitochondrial Dysfunction in Paraplegin-Deficient Mice."
It has long been recognized that the body’s cells are dependent on mitochondria for reactions that provide much of the energy they need to survive. Mitochondria are extremely complex organelles with numerous interacting components. Paramount among these are the enzymes that keep the energy-producing reactions flowing quickly and efficiently. Just as with the parts of a human-made machine, improperly manufactured proteins can negatively impact energy production by mitochondria.

Dr. Elena Rugarli and her team at National Neurological Institute in Milan, Italy, are conducting a UMDF-funded study of an enzyme (paraplegin) that normally removes defective mitochondrial proteins before they can hamper energy metabolism. She is using paraplegin-deficient mice as a model to investigate the human condition known as hereditary spastic paraplegia, characterizing the role that abnormal mitochondrial protein synthesis plays in this neurodegenerative disease. Gaining an understanding of the normal functions of paraplegin in mitochondria and the specific consequences of its loss is a necessary step towards developing effective treatments for this incurable disease.

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Mair Churchill, PhD
University of Colorado Health Sciences Center

2005 Research Award: $116,133

"Molecular Basis of Mitochondrial Gene Regulation"
Many of us have an understanding of the significance of the genetic code and how it contains the directions, in the form of DNA, for making the cells of the human body. The DNA code is "transcribed" into a related molecule known as messenger RNA, which is then "translated" into the proteins that make up the organelles. These organelles are the functional units of the cell and their amino acid structures can ultimately be traced back to the DNA genetic code. If you’ve looked through a microscope in a biology course to see the nucleus of a cell, you may have been told that this is where the DNA resides. It is true that the instructions for manufacturing the cell’s organelles are to be found in the nucleus. There is one organelle, however, for which this is only partially correct. Mitochondria, the primary sites of energy metabolism in the cells, actually contain some of the DNA that is necessary for their own formation. The process whereby the DNA stored in both the nucleus and the mitochondria are used to make mitochondrial proteins is extremely complex.

Dr. Mair Churchill and her team at the University of Colorado Health Sciences Center are conducting UMDF-funded research to better understand the transcription of mitochondrial DNA (mtDNA) into messenger RNA. This process of always "rewriting" the DNA code into an RNA code before it is used to makes proteins assures that the original message remains unchanged. Transcription is a complicated procedure that is under the control of specific regulatory molecules known as transcription factors. The mitochondria have their own set of these to assure that mtDNA is properly transcribed into messenger RNA. Dr. Mair is using X-ray crystallography methods to determine the exact molecular structures of these mitochondrial transcription factors. Being able to accurately describe their structure will provide insight into how they operate. This will, in turn, also improve our understanding of mitochondrial diseases that are caused by abnormal transcription factor activity.

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Linda Spremulli, PhD
University of North Carolina, Chapel Hill

2005 Research Award: $110,980 

"Biochemical and structural studies on mitochondrial disease mutations in methionyl-tRNA."
By now, many people are aware that their DNA contains the genetic information needed for the cells in their body to function normally. The DNA is actually a code that spells out the structure of the cells’ proteins and most of it is housed inside the nucleus. Mitochondria are the primary source of ATP, the high-energy molecule used by cells to carry out a variety of activities. An additional level of complexity was added to the understanding of mitochondrial function when it was discovered that mitochondria also contain DNA. How this organelle is produced when the genetic instructions for its manufacture are located in two separate cell compartments makes for a complex story. The DNA code is "transcribed" into a related molecule known as messenger RNA, which is then "translated" into the proteins that make up the organelles. Translation involves the delivery of specific amino acids to the messenger RNA template so that the correct protein can be manufactured. It is the job of the transfer RNAs (tRNA) to deliver the amino acids as they are required. Because mitochondrial tRNA is made from the genes in mitochondrial DNA, mutations in those genes can be inherited. Such inherited mutations can result in debilitating, and sometimes fatal mitochondrial disease.

Dr. Linda Spremulli and her team at the University of North Carolina, Chapel Hill, will investigate mutations that cause "mis-folding" of a mitochondrial transfer RNA that is linked to specific defects in the function of mitochondria. If a specific tRNA is unable to function properly, then protein synthesis will be compromised. This research will provide fundamental insights into how a malfunctioning transfer RNA originating inside mitochondria can cause deficits in mitochondrial energy production.

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Richard H. Haas, MB, BChir
University of California, San Diego

2005 Research Award: $109,991

"Diagnostic Utility of DHPLC in Mitochondrial Disease"
Some mitochondrial disease patients suffer through numerous misdiagnoses before finally learning of the true cause of their illness. This is not surprising since fatigue, muscle-weakness, gastrointestinal upset and other symptoms resulting from impaired mitochondrial function could also result from a number of other diseases. While a reduced ability to provide a constant flow of energy to the cells in the form of ATP is the common underlying pathology of mitochondrial illnesses, this basic defect can manifest itself in a variety of ways. Since effective medical practice involves achieving an accurate differential diagnosis, being able to either identify or rule out mitochondrial disease through a simple, straightforward test would be immensely helpful.

Dr. Richard H. Haas and his team at the University of California, San Diego, are conducting UMDF-funded research that will evaluate the potential of one such test. They are using blood and saliva samples from patients to develop simple, more reliable methods for detecting mitochondrial DNA mutations that can lead to disease. The long-term goal of this research is to develop standard procedures for rapid screening and diagnosis of mitochondrial disease.

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Luca Scorrano, MD, PhD
Dulbecco-Telethon Institute, Italy

2005 Research Award: $94,000 

"Role of mitofusin-2, a mitochondria-shaping protein mutated in Charcot-Marie-Tooth IIa, in controlling mitochondrial function and apoptosis."
A typical cell contains eight or so different kinds of organelles with specific functions that contribute to its growth and maintenance. Mitochondria are the organelles responsible for providing energy for the many things that a cell must do to survive. They must be present in sufficient quantities at all times and can be synthesized inside the cell as needed. Instead of being made from scratch, new mitochondria develop from pre-existing ones, with constant replenishing of the necessary proteins as directed by DNA in both the nuclei and in the mitochondria themselves. Interestingly, at any point in time variable numbers of mitochondria may fuse together to form elongated structures, or longer structures may fragment into the smaller bean-shaped structures often depicted in biology textbooks. Previous studies have demonstrated that the ability to fuse is important to maintaining normal mitochondrial function and that fusion is a highly regulated activity.

Dr. Luca Scorrano and his research team at the Venetian Institute of Molecular Medicine in Italy are studying abnormalities in mitochondrial shape and function that result from a mutated fusion-regulator called mitofusin 2. This protein is important in the maintenance of mitochondrial shape and a defect in it is associated with Charcot-Marie-Tooth IIa disorder, a type of muscular dystrophy. The importance of this research lies in the insights that will be gained into the role of mitofusin 2 in maintaining normal mitochondria and how disease results from its mutated form.

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Jan-Willem Taanman, PhD
University College London, UK

2005 Research Award: $86,455 

"The assembly pathway of human cytochrome-c oxidase studied with RNA interference"
Adenosine triphosphate (ATP) is the energy source used by the human body for all of its life processes. It took several decades during the first half of the 20th century to figure out how the potential energy in food is carefully transformed to ATP for the use of individual cells. Part of the challenge had to do with understanding the complexities of the mitochondrial enzymes and the contributions that each makes to the overall energy-transformation process. The cytochromes, vital components of this mitochondrial respiratory enzyme system, were first identified in the 1920’s but how they are manufactured inside the mitochondria has still not been fully worked out.

Dr. Jan-Willem Taanman and his team at the University College of London, U.K. are conducting UMDF-funded research to characterize in greater detail the assembly pathway of cytochrome-c oxidase, an important enzyme involved in mitochondrial ATP synthesis. This research will lead to detailed fundamental information concerning how a crucial mitochondrial enzyme is assembled and provide insight into mitochondrial diseases that result from its malfunction.

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Tal Mia Lewin, PhD
University of North Carolina, Chapel Hill

2005 Research Award: $70,525

"Barth Syndrome: A Mitochondrial Disease with Insights into Cardiolipin Synthesis"
Barth syndrome is a rare genetic disorder that causes generalized muscle weakness, affecting both the muscle of the heart and of the musculoskeletal system. It is a severe disease that also has a negative impact on a specific population of immune system cells. Previous studies have shown that the muscle mitochondria in Barth patients are deficient in cardiolipin, a compound that is an essential component of the mitochondrial inner membrane. Production of the ATP needed for cell activity is compromised in the absence of normal cardiolipin levels.

Dr. Tal Mia Lewin and her team at the University of North Carolina, Chapel Hill are conducting UMDF-funded research to investigate the role played by a defective enzyme in Barth syndrome that normally regulates the synthesis of cardiolipin. They are using cell cultures to identify the detailed steps leading to the manufacture of this important molecule. Investigating the fundamental pathology of this fatal mitochondrial disease can set the stage for development of effective treatments. Although Barth’s is a rare disease, this research should provide insight into the disease mechanisms of other mitochondrial disorders as well.

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David C. Chan, MD, PhD
California Institute of Technology

2004 Research Award: $128,000

"Understanding the role of mitochondrial fusion in mitochondrial myopathies"
Dr. Chan’s lab will investigate the formation of cell organelles called mitochondria that are responsible for energy production in human cells. Human cells contain several kinds of organelles, each of which has specific jobs. Mitochondria are especially complex machines made from a variety of proteins. The genetic code for the manufacture of most of these proteins resides in the DNA of the cell’s nucleus. Mitochondria are unique among organelles, however, because the nucleus does not contain all of the genetic information needed to make them. Some of this information resides in the mitochondria’s own DNA.

Dr. Chan’s lab will breed lab mice that have undergone tissue-specific deactivation of factors needed for mitochondrial fusion and will study the effects that this has on skeletal and cardiac myopathies. Instead of being made from scratch, new mitochondria always develop from pre-existing ones, with constant replenishing of the necessary proteins as directed by both the nuclear DNA and mitochondrial DNA. Interestingly, at any point in time variable numbers of mitochondria may fuse together to form elongated structures, or longer structures may fragment into the smaller bean-shaped structures often depicted in biology textbooks. The rate at which these events, division and fusion, occur is under the control of a specific group of enzymes called GTPases that reside in mitochondria. Some mitochondrial diseases are caused by defective mtDNA and fusion may actually protect the mitochondria from following the incorrect code of defective mtDNA.

Dr. Chan plans to investigate the role of three GTPases required for the fusion of mammalian mitochondria. Previous research has shown that mouse embryos lacking these will die early in their development. He will produce lab mice that have undergone deactivation of the GTPases needed for mitochondrial fusion, but only in their skeletal muscle and heart muscle. Because human mitochondrial diseases often affect function of these two types of muscle, this research will provide insight into the role that fusion of mitochondria plays in progression of the disease.

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Miriam H. Meisler, PhD
University of Michigan

2004 Research Award: $108,305

"The nuclear-encoded gene OMI and mitochondrial disease"
Dr. Meisler has been conducting research with mice that possess a mutation causing abnormal mitochondrial function. Cells contain several kinds of machines called organelles, each of which has specific functions. The organelles under study in Dr. Meisler’s lab are the mitochondria, especially complex cell organelles that are responsible for energy production. The mutated mouse gene is called OMI and it causes the affected mice to develop a neuromuscular disease that results in severely uncoordinated movement and wasting of muscles. There are a number of diseases that cause degeneration of the nervous system and the muscles that it controls. The changes experienced by the mice with the mutated OMI gene are a direct result of the inability of their abnormal mitochondria to provide adequate energy.

Building upon her experience with this mouse disease model, Dr. Meisler has designed a study in which she will screen 300 patients with inherited mitochondrial disease for OMI mutations similar to those found in mice. Mutated DNA sequences that occur in the mitochondrial disease patients, but not in healthy controls, will be tested for their involvement in the development of mouse neurodegenerative disease. This will be accomplished by microinjection of the candidate DNA sequences into fertilized eggs that would normally develop into mice with the fatal disease.

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Volkmar Weissig,
Northeastern University, Dept. of Pharmaceutical Sciences

2004 Research Award: $99,360

"Development of a method for transforming mitochondria in living mammalian cells with exogenous DNA"
Dr. Weissig’s research group is interested in developing a means of replacing defective DNA sequences in mitochondria with the correct sequences that are required for normal function. Mitochondria are complex energy-producing cell organelles that are made from a variety of proteins. The genetic code for the manufacture of most of these proteins resides in the DNA of the cell’s nucleus. Mitochondria are unique among organelles in that the nucleus does not contain all of the genetic information needed to make them. Some of this information resides in the mitochondria’s own DNA. If a defect is present in the mitochondrial DNA (mtDNA), then it will keep the mitochondria from making the normal proteins needed for energy production. Many mitochondrial diseases due to mtDNA defects develop into serious, ultimately fatal neuromuscular disorders. Gene therapy as an approach to treatment of these diseases holds great promise.

Dr. Weissig plans to develop and perfect a mechanism for transporting normal DNA through the interior of cells to the mitochondria, with subsequent mitochondrial uptake of the DNA. Researchers are also developing gene therapy approaches to the treatment of diseases such as muscular dystrophy. This is more straightforward than treatment of mitochondrial disease because it is easier to insert the correct DNA sequence into the nucleus of cell. Extra challenges are associated with inserting the correct DNA sequence into mitochondria. Development of a reliable method for insertion of DNA into mitochondria would be a significant step in treatment of often fatal mitochondrial diseases.

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Vamsi K. Mootha
Broad Institute, Massachusetts Institute of Technology

2004 Research Award: $90,200

"Genomic Approaches to Human Cytochrome c Oxidase Deficiency"
Dr. Mootha’s research team will identify genes that code for factors responsible for directing the synthesis of an important component of the mitochondrion. Mitochondria are energy-producing organelles contained in most human cells. They possess a series of molecules that are called collectively the respiratory chain. The respiratory chain is the cellular site where most of the energy is derived from foods that we eat to synthesize the energy-rich molecule ATP. The complex known as "cytochrome c oxidase" is an important component of the respiratory chain. Various assembly factors are required to direct the manufacture of new cytochrome oxidase complexes in the mitochondria.

Researchers have identified five human assembly factor mutations that can lead to abnormally functioning cytochrome c oxidase and diminished production of ATP by the mitochondria. Dr. Mootha has recognized the need for a comprehensive inventory of the nuclear encoded genes that are required for proper cytochrome c oxidase assembly and function. Computer analysis of data made available by the Human Genome Project can be used to perform a search for all human DNA containing the genetic code for cytochrome c oxidase assembly factors. Once specific DNA sequences have been identified as potential candidates, the research team will validate these sequences through the use of RNA interference in cultured cells. This procedure prevents the cells from expressing that particular DNA sequence. Subsequent biochemical assays of these cells will measure any changes in mitochondrial function that resulted from blocking expression of the DNA. Information gained from this research will increase our understanding of normal mitochondria function, providing insights into what goes awry when mitochondrial disease develops.

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Joseph A. Garcia, MD, PhD
University of Texas SW Medical Center at Dallas

2004 Research Award: $88,852

"The hypoxia sensing transcription factor EPAS1/HIF-2a is a novel mitochondrial disease candidate in mice and man"
Dr. Garcia’s lab will screen cells obtained from children with mitochondrial disease for the presence of an abnormal protein that has lost the ability to stimulate the production of anti-oxidant enzymes. Mitochondria are energy-producing organelles contained in most human cells. They contain a series of molecules that collectively are called the respiratory chain. The respiratory chain is the cellular site where most of the energy is derived from foods that we eat. Adequate amounts of oxygen must be available for the chain to function properly and produce the energy-rich molecule ATP. While we might consider only the benefits of getting enough oxygen as we take each breath, most of us are not aware of the potential problems if the oxygen is not quickly used in the mitochondria for energy production. Mitochondrial disease results from a variety of problems with the energy-producing machinery of the patient’s mitochondria. Abnormal mitochondria are unable to reliably and efficiently produce energy and will not use all of the oxygen that is available to them. The unused oxygen may accumulate to the point where it could begin to damage the mitochondria. This is because the presence of excess oxygen in the cells leads to the production of a variety of chemicals known collectively as reactive oxygen species (ROS). Free radicals and other types of ROS can remove electrons from the atoms that make up our cells. Extensive exposure to ROS can lead to oxidative stress, causing extensive damage to any tissues made of cells that contain abnormal mitochondria.

Cells possess several means of protecting themselves against damage from ROS, including antioxidant enzymes. The synthesis of these protective enzymes is regulated by the cellular protein EPAS1/HIF-2a (EH), which responds to oxidative stress by activating specific genes. Upon receiving a signal from EH, these genes direct the production of antioxidant enzymes that will then minimize ROS damage. Dr. Garcia’s lab will screen cells that have been collected from 200 children with mitochondrial disease for the presence of mutated genes that provide the code for the EH regulatory protein. They will then insert these coding errors into cultured cells in the lab to determine the specific nature of the dysfunction associated with the protein.

Dr. Garcia will also work with mice to further understand the regulation of antioxidant enzymes. Loss of EH in mice results in increased levels of ROS and causes them to develop a condition that is similar to mitochondrial disease in humans. This similarity provides an excellent animal model for study of the disease. Dr. Garcia will work both with mice that lack EH and with normal control mice, comparing the capacity of their skeletal muscle cells to produce the energy-containing molecule ATP.

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Stefan Strack
University of Iowa, Carver College of Medicine

2004 Research Award: $88,000

"Protein phosphatase 2A in mitochondrial function and disease"
Dr. Strack’s laboratory is interested in conducting basic research that could lead to the development of new treatments for mitochondrial disease. Mitochondria are energy-producing organelles contained in most human cells and can cause specific disease conditions when they are damaged. Dr. Strack will develop methods of blocking the activity of a mitochondrial regulatory protein that normally renders brain cells vulnerable to toxin-induced degeneration. His ultimate goal is to find a means of rescuing the cells from the effects of the toxins. His research group is interested in neurodegenerative diseases such as Huntington’s chorea and Parkinson’s disease.

Previous research identified a regulatory protein that is localized to mitochondria in cells of the brain. The protein renders cultured brain cells vulnerable to damage by toxins. If the protein is inhibited, then resistance to the toxins is conferred upon the cells. In his proposal, Dr. Strack suggested that development of drugs against the protein may provide an effective treatment for mitochondrial disease. He will use genetically-engineered viruses to place inhibitors specific to the protein into the brains of rats, followed by testing for resistance to neurotoxins that normally poison the brain. Finding effective means of rescuing the cells from the effects of the toxins could provide insight into prevention of the nervous system degeneration that is typical of some mitochondrial diseases.

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Brian Robinson
Hospital for Sick Children, Canada 

2004 Research Award: $44,000

"Drug development for the regulation of respiratory chain components in mitochondria"
Dr. Robinson’s research team will screen a large number of chemicals from a family of compounds that show promise for stimulating the production of mitochondrial respiratory proteins. Mitochondria are energy-producing organelles contained in most human cells. They possess a series of energy-producing molecules that collectively are called the respiratory chain and are responsible for synthesizing the energy-rich molecule ATP. Some mitochondrial diseases result from deficiencies of certain respiratory enzymes that are part of the chain. But these defects are rarely total, with some capacity for ATP production remaining. Dr. Robinson’s goal is to look for drugs that could be used to enhance whatever residual activity is present in abnormal mitochondria, so as to increase overall energy production in mitochondrial disease patients.

Prior discovery of two nuclear transcription factors that increase the rate at which mitochondrial respiratory chain proteins are synthesized has led to Dr. Robinson’s suggestion that drugs could be developed to stimulate the activity of these factors. Such stimulation is indeed what happens in response to exercise. Dr. Robinson has access to a large chemical library of heterocyclic compounds that may stimulate the transcription factors in the absence of exercise. He plans to conduct a systematic screening of these chemicals to identify ones that show promise for stimulating the production of mitochondrial respiratory proteins.

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Gregory M. Enns, MB, ChB
Stanford University, California

2004 Research Award: $34,179

"GSH levels, reactive oxygen species production, lipid peroxidation, products and mitochondrial membrane potential in patients with mitochondrial disease"
Dr. Enns' laboratory will look for correlations between the relative health status of mitochondrial disease patients and various factors associated with oxidative stress in the patients' cells. Mitochondria are energy-producing organelles contained in most human cells. They contain a series of molecules that collectively are called the respiratory chain. The respiratory chain is the cellular site where most of the energy is derived from foods that we eat.

Adequate amounts of oxygen must be available for the chain to function properly. While we might consider only the benefits of getting enough oxygen as we take a breath, most of us are not aware of the potential problems that occur if that oxygen is not quickly used in the mitochondria for energy production. Abnormal mitochondria are unable to reliably and efficiently produce energy and will not use all of the oxygen that is available to them.

The unused oxygen may accumulate to the point where it could begin to damage the mitochondria. This is because the presence of excess oxygen in the cells leads to the production of a variety of chemicals known collectively as reactive oxygen species (ROS). Free radicals and other types of ROS can remove electrons from the atoms that make up our cells. Extensive exposure to ROS can lead to oxidative stress, causing extensive damage to any tissues made of cells that contain abnormal mitochondria.

Cells have several ways to protect against damage from ROS, including the glutathione defense system. Dr. Enns' laboratory will study glutathione levels and oxidative stress in mitochondrial disease patients. Specifically, they will determine the degree of oxidative stress and levels of glutathione in patients when they are outwardly healthy and compare them to times when the patients are acutely ill. Dr. Enns will also measure the extent to which the mitochondria in these patients have been damaged. Measurements of oxidative stress could become a means of providing insights into the effectiveness of patient treatment. Another goal is to assess the potential of a compound known as N-acetylcysteine therapy to decrease the incidence of oxidative stress.

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Ramon Marti, PhD
Fundacio Institut Hospital Vall d'Hebron, Spain

2004 Research Award: $33,776

"Restoration of thymidine phosphorylase activity in MNGIE patients through platelets infusion"
Dr. Marti’s lab is investigating a rare disease that affects the digestive system. Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) is a disease of the mitochondria, cell organelles that are responsible for energy production. The severely disrupted digestive function associated with MNGIE typically leads to death in the early adult years. Mitochondria are one example of the complex machines, called organelles, that are found in the human cell. These organelles are made from a variety of proteins. The genetic code required for the manufacture of most these proteins resides in the DNA of the cell’s nucleus. Mitochondria are unique among organelles, however, in that the nucleus does not contain all of the genetic information needed to make them. Some of this information resides in the mitochondria’s own DNA. In order to replicate their own DNA, mitochondria require the activity of an enzyme called thymidine phosphorylase. Unfortunately, a mutation in the gene that provides the genetic code for thymidine phosphorylase causes the production of an abnormal version of the enzyme. The resulting buildup of thymidine, which normally is phosphorylated during the replication of mitochondrial DNA, has a toxic effect on mitochondria and over time will severely diminish the energy-producing capacity of the cells in which these damaged organelles reside.

Platelets, blood cells involved in clotting, contain a number of enzymes and biologically active compounds and are especially rich in thymidine phosphorylase. Dr. Marti plans to infuse MNGIE patients with platelets obtained from healthy blood donors. The expected outcome is that the plasma levels of toxic thymidine in the MNGIE patients will be significantly reduced. Patients will receive platelet infusions every week for one month, during which plasma thymidine levels will be measured and any changes in clinical status will be monitored.

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Immo Scheffler, PhD
University of California, San Diego

2003 Research Award: $100,000

"Application of RNA interference in the study of NADH-ubiquinone oxidoreductase (complex I) assembly in mammalian mitochondria"
Small interfering RNAs (siRNAs) can nowadays be readily produced in many cells including mammalian cells. RNA interference (RNAi) has become a novel and powerful method to knock down the expression of specific genes in such cells, an alternative to the classical approach utilizing mutations. With the complete knowledge of many of the genomes, any gene can in principle become a target, and hence any protein of interest can be eliminated from a cell to study the physiological and pathological consequences.

Our interest is focused on a large complex in the inner mitochondrial membrane that is essential for respiration (complex I, NADH-quinone oxidoreductase). The complex has a total of 45 subunits, of which 38 are encoded by nuclear genes, and 7 are encoded by mitochondrial genes. Thus, 38 genes are potentially subject to manipulation by RNAi. A fundamental issue is that 14 orthologous proteins in bacteria can perform a very similar function, and the challenge is to elucidate the role of over 30 additional subunits that have been added to the complex in mammalian mitochondria in the course of evolution. RNAi-based technology will for the first time permit a systematic examination of the role of each of these nuclear-encoded subunits in the assembly, stability, activity and regulation of complex I. The insights gained can be applied to the diagnosis and understanding of mitochondrial diseases, particularly the growing class of such diseases resulting from a (partial) complex I deficiency. Partial deficiencies can arise either from a reduced specific activity of the complex or from lower levels due to assembly defects. In the longer term, the technology can be adapted to perform such experiments in whole animals such as a mouse, with the prospect of developing good animal models for the study of the broader physiological and pathological aspects of these diseases.

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Mikhail Alexeyev, PhD
University of South Alabama

2003 Research Award: $100,000

"Selective Elimination of Defective Mitochondrial Genomes as an Approach to the Reversal of NARP and MILS Syndromes, Heritable Mitochondrial Disorders"
The NARP and MILS syndromes are devastating disorders caused by T8993G mutation in mitochondrial DNA. There exists a compelling evidence that this mutation is the most commonly identified mtDNA mutation in children. Clinically, NARP is characterized by sensory neuropathy, cerebellar ataxia, retinitis pigmentosa, dementia, seizures and developmental delay. MILS, on the other hand, is usually associated with more severe manifestations and early onset (4 to 5 months). Patients with NARP and MILS syndromes are typically heteroplasmic, which means that their cells have both normal and mutant mitochondrial DNA. The difference in the clinical severity is largely due to the mutant load (i.e. % of mutant mtDNAs in every given cell). The life expectancy and quality of life in patients is believed to inversely correlate with mutant load. Thus, patients with MILS have very high mutant loads, typically >90% mutant mtDNA. NARP is usually associated with intermediate mutant loads of 60 to 80%, and mutant loads less than 60% are generally asymptomatic. Therefore, it appears that lowerlng the mutant load below the 60% threshold should be sufficient to render both NARP and MILS patients asymptomatic.

Amazingly, two bacteria, Serratia marcescens and Xanthomonas malvacearum, produce enzymes that can selectively recognize and destroy mutant, but not normal mitochondrial genomes with 99.95% efficiency in a test tube. Therefore, the central idea of this application is to achieve therapeutic effect through targeting of bacterial enzymes to mitochondria. If successful, this approach may allow, for the first time ever, to revert, rather than to just treat, the underlying mitochondrial disease. Our calculations indicate that even if bacterial enzymes will be only 1 % as efficient in mitochondria as they are in a test tube, they still should be able to bring mutant DNA content down below 60% threshold in a patient with 93% mutant mitochondrial DNA. Preliminary data indicate viability of the proposed approach. Thus, the focus of this proposal is to further experimental procedures to target therapeutic enzymes to mitochondria.

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Matthew Freeman, PhD
Laboratory of Molecular Biology - M.R.C., UK

2003 Research Award: $90,000

"Role of Rhomboid Proteolysis in Optic Atrophy"
Dominant optic atrophy is a genetic disease that causes early childhood blindness with prevalence as high as 1: 10,000 and is the result of defects in a protein called OPA1. OPA1 is a protein that is responsible for maintaining the proper structure and function of the mitochondria, a vital part of the every cell. For OPA1 to function it needs to be cut at the correct place and the correct time by another protein in the cell. We have recently characterized the protein, called Rhomboid, which cuts OPA. We believe that this may be an important step in describing the molecular mechanism of this disease. By using the mouse as a model organism we hope to determine the link between OP A and Rhomboid regulation of mitochondrial function and human optic atrophy.

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Koji Okamoto, PhD
University of Utah

2003 Research Award: $83,400

"Molecular Basis of Mitochondrial Membrane Dynamics: a New Paradigm of Human Disease"
Mitochondria are dynamic organelles that change size and shape in order to optimize their energy production and supply for cellular functions. Loss of normal mitochondrial morphology results in a variety of human diseases including neurological disorders and some types of cancer. Changes in mitochondrial morphology are also associated with programmed cell death and aging in humans. Although recent findings suggest that deregulation and loss of mitochondrial fusion may contribute to defects associated with mitochondrial diseases, very little is known about the mechanisms that regulate mitochondrial fusion. To understand pathogenesis leading to such diseases, regulatory molecules for mitochondrial fusion must be identified and characterized.

The transmembrane GTPase, called Fzo1 in yeast, is essential for mitochondrial fusion in both yeast and humans. Recent analyses in yeast suggested that GTP hydrolysis by Fzo1 is required for mitochondrial fusion. Regardless of which fusion step requires GTP hydrolysis by Fzo1, it seems very likely that a GTPase-activating protein (GAP) plays a crucial role in modulating mitochondrial fusion. To understand how the GTP-driven molecular device regulates mitochondrial membrane fusion, we will identify a GAP that stimulates GTP hydrolysis by Fzo1 in yeast. We will then analyze this GAP by a combination of biochemical, cell biological and genetic approaches. Moreover, we will investigate the GAP-mediated stimulation of the Fzo1 GTPase activity in vitro.

Since Fzo1 is an evolutionarily conserved GTPase, it is also likely that yeast Fzo1 GAP has a human homologue. What we learn about the mechanism of mitochondrial fusion in budding yeast will be directly relevant to the mechanism of mitochondrial fusion in humans. The proposed project will help determine how the Fzo1 GTPase cycle controls mitochondrial fusion, allowing researchers to investigate the role of regulated fusion and mitochondrial fragmentation during apoptosis. In addition, our research will provide insight into molecular bases for mitochondrial fusion that could contribute to the future development of gene therapies based on inter- mitochondrial complementation. Finally, this study will promote establishment of animal models for understanding pathogenesis of human diseases associated with mitochondrial fusion defects.

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Bernard Lemire, PhD
University of Alberta, Canada

2003 Research Award: $76,780

"The Use of the Yeast CYB2 Gene As Therapy for Complex I Mutations in a C. elegans Model System"
The mitochondrial respiratory chain (MRC) is the major source of energy for most cells and tissues in the human body. It captures energy from the food we eat by catalyzing the transfer of electrons from NADH, which is derived from that food, to the oxygen we breathe. When MRC function is impaired, NADH accumulates and is diverted towards the formation of lactic acid resulting in lactic acidosis. This condition can produce malaise, weakness, exercise intolerance, and vomiting. It may also contribute to the long-term progression of and developmental delays associated with mitochondrial diseases by modulating the expression of genes related to energy metabolism.

We propose to investigate the use of a yeast enzyme called cytochrome b2 that will directly act to reduce the levels of both lactic acid and NADH. We have isolated and studied a series of complex I MRC mutants that mimic known human mutations in the nematode, Caenorhabditis elegans. The nematode is a very simple animal with an MRC that closely resembles its human counterpart and serves as a sophisticated genetic model system. We will introduce the DNA encoding cytochrome b2 into each of the mutants and evaluate how the yeast protein affects animal fitness as measured by fertility , motility , lifespan, and levels of lactic acid. These results will address the contribution of lactic acidosis to mitochondrial diseases and may lead to the development of a new therapy. Our results will lead to a better understanding of the fundamental biological processes surrounding mitochondrial energy production in normal and in disease states.

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Giovanni Manfredi, MD, PhD
Weill Medical College of Cornell University, New York

2003 Research Award: $50,000

"MtDNA complementation and recombination in mitochondrial disorders"
Mutations in the mitochondrial DNA (mtDNA) cause mitochondrial diseases. MtDNA is maternally inherited. Thus, mutations in the mtDNA typically result in family pedigrees exhibiting maternal inheritance, i.e. the disease should pass only through females, and essentially all the children inherit the mtDNA mutation. However, despite this simple way of inheritance, the manifestation of mtDNA-related diseases may be very variable even within the same family.

Often, in patients with mitochondrial diseases associated with mtDNA mutations, normal and mutated mtDNAs may coexist within cells and tissues, a condition known as heteroplasmy. An important, but still not well understood, concept in the genetics of mitochondrial diseases is that of complementation in heteroplasmic cells. It is the subject of debate whether in human mitochondria functional complementation between mtDNA molecules is an efficient process or even whether it occurs at all. This is a very important question because if mutant mtDNA molecules function as independent units, unable to interact across different organelles, the protective effect of normal molecules coexisting with the mutated ones (heteroplasmy) would be rather limited.

Moreover, the consequences of randomly occurring new mtDNA mutations, for example acquired during aging, would be more severe in the absence of efficient complementation between normal and mutated mtDNA molecules. This is especially relevant for patients that already harbor an inherited pathogenic mtDNA mutation, in whom the occurrence of acquired mutations may precipitate the clinical phenotype. To address these issues, we have generated a hybrid cell culture model from the fusion of two human cell lines with identical nuclear DNA, but each with a distinct mutation in the mtDNA.

This model allows us to study complementation among mutated mtDNAs in a controlled system. Our goal is to better understand the mechanisms underlying mitochondrial complementation, because we believe that this may contribute to the identification of novel tools for the treatment of these diseases.

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Philip Schwartz, PhD
Children's Hospital of Orange County, California

2002 Research Award: $81,574

"Electrophysiologic Properties of Neural Stem Cells from Patients with Mitochondrial Disease"
One of the more prevalent and debilitating features of mitochondrial disease is seizures. Seizures result from uncontrolled electrical activity within the brain. This altered electrical activity may result from changes in any number of basic properties of neurons, the functional cellular units of the brain. Unfortunately, our scientific understanding of the electrical changes in these cells is very incomplete as there has been no way to study neurons with mitochondrial disease while these cells are alive and functioning. Our current understanding of these changes comes only from inferences made from studying EEGs, the responses of patients to various drugs, and the brains of patients that have died with mitochondrial disease.

Recently, my colleagues and I have shown that we can harvest living neural stem cells from the post-mortem human brain, that is from the brain after death of the individual. The same cannot be done for neurons as neurons die very rapidly after death of the individual. Neural stem cells do not. Neural stem cells are immature brain cells that can, in the laboratory, be coaxed into becoming mature neurons. By harvesting neural stem cells from patients that have died with mitochondrial disease and growing these cells in the laboratory, we have produced a living, functioning, brain cell preparation with which we can now study the electrical changes associated with these cells in these same patients.

In the work described in this proposal, therefore, neural stem cells will be harvested from patients with mitochondrial disease and from patients with no neurologic disease (for comparison). This will be done in collaboration with the National Human Neural Stem Cell Resource which I direct. The cells will be grown up in my laboratory and coaxed into becoming electrically active neurons. The electrical properties of these cells, under a variety of conditions, will then be studied in detail. Importantly, we will study not only the basic electrical properties of these cells but also their responses to certain nutrients and drugs so that we may identify better potential anti-seizure treatments for patients with mitochondrial disease.

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Yidong Bai, PhD
University of Texas Health Science Center at San Antonio

2002 Research Award: $66,000

"Exploiting the potential of yeast NDI1 gene in the therapy of diseases linked with mtDNA"
The mitochondrial complex I, is the largest and least understood component of the energy producing system, consisting at least 43 subunits. Mutations in genes encoding subunits of complex I are associated with diseases, in particular the various forms of Leber's hereditary optic neuropathy (LHON) that cause blindness. Impaired complex I activities have also been reported to be related to other neurodegenerative diseases and aging. The ability to repair mutations in mitochondrial complex I via gene therapy holds the promise of treatment of mitochondrial diseases such as LHON. In contrast to the multisubunit complex I enzyme found in mammalian cells, the corresponding enzyme in yeast, NDI1, is a single polypeptide. Previously, we showed that the yeast NDI1 is active in a human cell line with an artificial nonsense mutation in mitochondrial DNA (mtDNA) ND4 gene. In this study we plan to further determine if the NDI1 gene can rescue mitochondrial function in cells carrying an mtDNA mutation that causes LHON in humans. More detail analysis of the activities of NDI1 gene in mammalian cells will also be performed. Finally we will initiate the production of transgenic mice expressing a complex I mutation transfected with NDI1 gene. The success of this study will significantly advance our endeavor to use gene therapy for the treatment of mitochondrial disease caused by complex I deficiency.

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Tanja Taivassalo, PhD
Institute for Exercise & Environmental Medicine, Texas

2002 Research Award: $61,389

"Exercise-induced mitochondrial gene shifting: Resistance training as a therapy for sporadic mtDNA mutations"
A clinical research trial is proposed to determine the efficacy of a novel approach to therapy for selective patients with mitochondrial disorders. The approach is based on the concept of "mitochondrial gene shifting" through exercise training, where resistance exercise is used to shift a patient's own normal mitochondrial genes from muscle precursor cells into their existing skeletal muscle which contains high levels of abnormal mitochondria and mitochondrial DNA. This has the potential to reverse the accumulation of abnormal mitochondrial DNA that occurs over time within skeletal muscle, restore a more normal function of muscle mitochondria, and thereby increase the energy-generating capacity of the cell. Also, normal adaptive increases in muscle strength associated with resistance training would be expected to further improve overall functional and physical capacity in patients, particularly in those demonstrating muscle weakness in addition to decreased muscle endurance. Due to the non-invasive nature and practicality of this approach, we believe resistance exercise training offers an imminent realistic advance in therapy and will be of functional significance in the quality of life of patients affected with mitochondrial myopathies.

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Jose Hernandez-Yago, PhD
Institute for Cell Research, Spain

2002 Research Award: $41,085

"Transport diseases in mitochondria: Full screening of DNA alterations in human genes encoding TOMM and TIMM complexes in patients with mitochondrial diseases"
The term "mitochondrial diseases" encompasses a heterogeneous group of disorders in which a primary mitochondrial dysfunction is suspected or proven by morphologic, genetic or biochemical criteria. The dual genetic control of mitochondrial proteins and the complex mechanisms needed for their synthesis, transport and correct assembly explain why a variety of genetic errors can cause mitochondrial diseases.

Many mitochondrial diseases suggest dysfunction in the transport processes of mitochondrial proteins from the cytosol to their mitochondrial compartments. We propose in this project the complete screening of the human genes encoding all the subunits of the mitochondrial protein transport machinery (TOMM and TIMM complexes) in patients with mitochondrial diseases as well as in the general population. The finding that the Mohr-Tranebjaerg syndrome-a mitochondrial disease that is a recessive degenerative syndrome characterized by postlingual progressive sensorineural deafness as the first presenting symptom in early childhood, followed by progressive dystonia, spasticity, dysphagia, mental deterioration, paranoia and cortical blindness- is due to defects in the hTimm8a gene, is a significant example of the severe consequences resulting from mitochondrial protein transport dysfunction.

In our laboratory we use a high throughput technology to screen DNA alterations that makes viable the screening of the ca. 100 exons of the approximately 15 subunits of TOMM and TIMM complexes. It is noteworthy that these exons are short enough to be amplified as a whole, using flanking intron primers.

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Min-Xin Guan, PhD
Children's Hospital Medical Center, Ohio

2001 Research Award: $33,000

"Biochemical Basis for Maternally Inherited Deafness"
Hearing loss is the most frequent sensory disorder. One in 1000 children is born deaf, an equal number lose their hearing by adulthood and half the population experience significant hearing impairment by the age of 65 years. Deafness can be due to genetic or environmental causes or a combination of both. Despite the recent progress in molecular characterization of deafness, the biochemical and molecular pathogenic mechanisms underlying the maternally inherited deafness remain poorly understood.

Recent results of genetic studies showed that an African-American fanilly with maternally inherited nonsyndromic hearing loss have been associated with the mitochondrial T7 511 C mutation in the tRNA Ser(UCN) gene, which is commonly related to deafness. To elucidate the pathogenic mechanism of this mutation, we have constructed a disease cell model by transferring mitochondria from lymphoblastoid cell lines derived from deaf individuals with mtDNA mutations or from controls lacking mutations, into human mtDNA-less (pO) cells. This application proposes two aims: 1). These transmitochondrial cell lines will be analyzed for the presence and severity of mitochondrial dysfunction associated with mtDNA mutations. 2). To study if over-expression of human mitochondrial Seryl-tRNA synthetase in these transmitochondrial cell lines can suppress the biochemical phenotype associated with T7511C mutation.

Success of this project will enhance a better understanding of pathogenic mechanisms of matemally inherited deafness, lead to the future therapies directed toward specific underlying abnormalities, and the development of animal models to test them.

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Brian Robinson, PhD
The Hospital for Sick Children, Canada

2001 Research Award: $15,000

"Efficacy of prenatal diagnosis of mitochondrial diseases."
Prenatal diagnosis of mitochondrial diseases by examination of biochemical metabolite or enzyme profiles is undoubtedly possible, at least for some types of disease. We propose that the biochemical abnormalities in fibroblasts of affected children can be documented and used to carry out prenatal diagnosis in amniotic cell cultures. While we have been collecting data from prenatal diagnosis of nuclear-encoded mitochondrial diseases for a number of years, this data set is far from complete. We have had one year of funding from UMDF to continue this project, one more year of funding should allow us to complete our studies and make some conclusions. This project will be funded to see if this hypothesis can be verified for a number of mitochondrial diseases to include cytochrome oxidase deficiency, complex I deficiency and pyruvate dehydrogenase complex deficiency.

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Edwin Kirk, MD
Sydney Children's Hospital, Australia

2001 Research Award: $12,000

"Complex I: The role of nuclear genes in disorders of childhood due to mitochondrial Complex I deficiency."
Complex I is a collection of proteins located in the mitochondria of the cell. These proteins work together to drive the first steps in the production of energy. Children with Complex I deficiency, often suffer devastating consequences such as deterioration of brain, muscle and other organ function.

They often die in the first years of life. Complex I deficiency is a genetic disorder, but in most cases, the underlying genetic fault is not known. There are at least 42 different protein building blocks which combine to make Complex I, each of which is coded for by a different gene. Because there are so many possible genes involved, finding the gene fault in a child with Complex I deficiency has been a very difficult task. Recently, however, a group of scientists in the Netherlands looked at 8 complex I genes which seem to be particularly important (because they have changed the least during evolution).

They found gene faults (mutations) in a total of 5 out of 20 families. We have samples from 40 children with proven complex I deficiency available for this study. This is a unique resource and represents all children in Australia with confirmed Complex I deficiency. We plan to search for mutations in the 8 important genes mentioned above, in the 40 families for whom we have DNA samples. If a mutation is found in a family, then the child's medical history will be reviewed. We will seek to identify patterns in the types of problems experienced by children with faults in each gene, to try to guide searches for faulty genes in future patients. If the faulty gene in a family is identified, it will enable us to provide answers for the family about the cause of the condition and how it is inherited in their family. For some, this information may also provide the option of testing future pregnancies to find out whether the baby is affected.

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Dikoma Shungu, PhD
Columbia University, New York

2000 Research Award: $36,719

"Quantitative In Vivo 1H Magnetic Resonance Spectroscopic Imaging of Cerebral Lactate as a Screening Test for Mitochondrial Disorders"
This research will seek to develop a test for screening patients suspected of having a mitochondrial disease. The test will be noninvasive in that it will be based on using magnetic resonance spectroscopic imaging (MRSI)—a technique that is nearly identical to standard MRI, except that it can measure levels of various biochemicals in the human body—to precisely quantify levels of lactic acid in the cerebrospinal fluid (CSF) of suspected patients. We believe this to be worthwhile because studies we have conducted over the past 5 years on many patients with mitochondrial diseases have shown that a large proportion of these patients have measurable levels of lactic acid in their CSF, whereas control individuals did not. If further refined, this technique for measuring lactate in the CSF may prove to be a very useful tool for screening patients with mitochondrial diseases, and helping assess the degree of severity of the disease and its progression in future clinical trials of promising therapies.

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George Perry, MD
Case Western Reserve University, Ohio

2000 Research Award: $18,281

"Is oxidative damage a result of metabolic abnormalities in Alzheimer disease?"
Evidence continues to accumulate that increased oxidative damage is important in Alzheimer disease (AD) and may be the biochemical basis of the increased incidence of AD with aging. In published studies, we have shown oxidative damage affects every neuron in susceptible neuronal populations in cases of AD, but not in normal people. In unpublished studies, we show that the only abnormality of vulnerable neurons in AD is changes in mitochondria. This proposal will specifically investigate the idea that oxidative damage in AD begins as mitochondrial abnormalities. To address this issue, we will 1) determine the relationship between oxidative damage, mitochondrial abnormalities and the pathology of the disease; and 2) determine whether some of the relationships noted in Aim 1 for AD hold true in Down syndrome. Completion of these aims will help to define the relationship of oxidative damage to mitochondrial abnormalities.

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Cecilia Giulivi, PhD
University of Minnesota, Duluth

1999 Research Award: $31,137

"Characterization of Mitochondrial Nitric-Oxide Synthase"
Expansion of the field of mitochondrial disease can be anticipated in several directions. First, into age-related degenerative diseases; second, into areas connected with new concepts in mitochondrial biochemistry and physiology. Of great interest are thorough studies aiming at a better understanding of the processes underlying mitochondrial defects. In this context, the recent finding of an enzyme, i.e., mitochondrial nitric-oxide synthase, that generates nitric-oxide, well-known modulator of important biological processes such as immunity, vasodilation, and neurotransmission, offers a new perspective in mitochondrial bioenergetics. We reported that the production of nitric-oxide by mitochondria modulates the synthesis of ATP; this latter compound constitutes the currency that supports cellular work. Considering the important role that nitric-oxide might have on ATP generation, this project is aimed at identifying the nitric-oxide synthase in different tissues, providing basis for future studies that will address strategies for attempts at the treatment of mitochondrial diseases associated with a defective enzymatic expression.

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Brian Robinson, PhD
The Hospital for Sick Children, Canada

1999 Research Award: $8,863

"Efficacy of Prenatal Diagnosis of Mitochondrial Diseases"
Prenatal diagnosis of mitochondrial diseases by examination of biochemical metabolite or enzyme profiles is undoubtedly possible, at least for some types of disease. We propose that the biochemical abnormalities in fibroblasts of affected children can be documented and used to carry out prenatal diagnosis in amniotic cell cultures. While we have been collecting data from prenatal diagnosis of nuclear-encoded mitochondrial diseases for a number of years under a small program funded by the US March of Dimes, this data set is far from complete. This project will be funded to see if this hypothesis can be verified for a number of mitochondrial diseases to include cytochrome oxidase deficiency, Complex I deficiency and pyruvate dehydrogenase complex deficiency.

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John Shoffner, MD
Horizon Molecular Laboratory, Georgia

1998 Research Award: $30,000

"Gene mutations in Leigh's Disease"
Leigh’s disease is a disorder in which degeneration in the basal ganglia structures, brainstem, and