By Anthony J. Hickey, Ph.D.
If you were to ask a biological scientist (and many non-scientists for that matter) about mitochondria, most would give you the standard answer that mitochondria function as “the powerhouse of the cell.” But this statement fails to truly reflect the complexity, dynamics, and even elegance of these vital cellular organelles. Four NICHD experts explained through their own work how mitochondria are more than just cellular batteries during February’s NICHD Exchange meeting: “The many faces of mitochondria in health and disease—it’s not just about ATP."
Mitochondria are unique organelles in eukaryotic cells, both in structure and evolutionary history. They have their own genome, which replicates independently from the cell’s nuclear DNA. Current evolutionary theory regarding mitochondrial origin states that these organelles descend from a bacterium (most likely Rickettsia) that was engulfed by an ancient eukaryotic precursor cell. The mitochondria escaped digestion and flourished within its new host by providing large amounts of ATP, a high energy molecule used to “fuel” almost all biochemical processes. With time, the two cells developed an endosymbiotic relationship, where neither could live independently of the other, giving rise to what is believed to be the first eukaryotic cell.1,2
Dr. Tracey Rouault opened the meeting with her talk “Mitochondrial Energy Capture: A Complex Key to Mammalian Evolution and Health.” Mitochondria generate the bulk of the cell’s energy through the creation of an electro-chemical gradient that ultimately provides energy for the synthesis of ATP.1 Dr. Rouault and her colleagues asked where the energy to create the electro-chemical gradient comes from.
It turns out that iron-sulfur clusters within mitochondrial proteins are key. They allow for the low energy capture and transfer of electrons through the electron transport chain, a process required for electro-chemical gradient formation. The synthesis of these iron-sulfur complexes themselves is complex. It involves the gradual assembly of multiple proteins onto a scaffolding protein named ISCU. After synthesis, the iron-sulfur clusters are transferred from ISCU to recipient proteins with the help of multiple molecular chaperones and co-chaperones. Dr. Rouault and her team have identified three key amino acids (Leucine, Tyrosine, Arginine) that engage these co-chaperones to ensure clusters are transferred to the correct location.3
Dr. Rouault concluded her talk by discussing a recessive, hereditary muscle disease associated with iron-sulfur biogenesis. Symptoms include exercise intolerance, mitochondrial iron overload, and deficiencies in the production of specific iron-sulfur proteins. Individuals afflicted with this myopathy make little to no functional ISCU protein. DNA sequencing of patient samples identified a single nucleotide change within this gene that results in the abnormal splicing of the gene’s transcribed RNA product.4
Dr. Rouault’s laboratory entered into collaboration with a pharmaceutical company known to design customized antisense oligonucleotides that can bind target RNA molecules. They hoped to generate oligonucleotides capable of preventing abnormal splicing of the ISCU transcript. Their preliminary results indicate that antisense oligonucleotide therapy is a viable line of investigation for treatment of this rare disease.
During the next talk, Dr. Jennifer Lippincott-Schwartz elaborated on the unique structure and physiology of mitochondria and its important role in maintaining metabolic homeostasis of the cell and the organism. She seeks to understand how mitochondria help cells to survive under different environmental conditions, specifically starvation.
Dr. Lippincott Schwartz and her team demonstrated, using their renowned expertise in fluorescent microscopy, that starvation initiates a process in cells where fatty acids are trafficked from lipid droplets to mitochondria. Her lab elucidated the role of autophagy during this process, a mechanism employed by cells during starvation, where cellular organelles are broken down and recycled,5 which turns out to be critical for replenishing lipid droplet stores during starvation by transferring fatty acids to them from membranes.
Dr. Lippincott-Schwartz next asked how lipid droplets interact with mitochondria to initiate fatty acid transfer, which appears to be through direct contact. Under starvation conditions, mitochondria convert from being individual bacteria-like organelles into a network of fused interconnected tubules.5 The disruption of fusion leads to fragmented mitochondria and an uneven distribution of fragment-lipid droplet interaction. Some mitochondria receive little to no fatty acids, while others receiving fatty acids at toxic levels. This also causes a disruption in fatty acid transfer and subsequent oxidation, which is evident from an increase in lipid droplet size. Eventually these molecules begin to accumulate in the cytoplasm at dangerous levels, requiring the cells to secrete them into the extracellular environment where they can be taken in by neighboring cells.
Dr. Lippincott-Schwartz concluded her talk discussing the ramifications such events have, not only on the physiology of the cell, but for the entire body as well. Some mitochondrial diseases are associated with high serum levels of fatty acids, and her lab’s findings add to our understanding of the adverse effects associated with diseases such as obesity and diabetes.
Dr. Danuta Krotoski, the third presenter of the afternoon, focused on the clinical aspects of mitochondrial dysfunction. Mitochondrial diseases have a large range of presentations, including neurological disorders, stroke, ataxia, epilepsy, migraines, exercise intolerance, and myopathy. Thus far, 100 different mitochondrial disease genes have been identified, but the number is sure to increase.
Dr. Krotoski’s talk, “Mitochondrial disorders: a tale of two genomes,” introduced the dilemma that mitochondrial disease can result from mutations in either mitochondrial or nuclear genome. While 42 mitochondrial diseases are known, she suggested the potential for more due to the difficulty in identifying and diagnosing these diseases, compounded by the multiple biochemical pathways that mitochondria influence and continuous crosstalk between the mitochondria and nucleus.
To illustrate the complexity of diagnosing and treating mitochondrial disease, Dr. Krotoski discussed three types of mitochondrial disorders: MELAS, Leigh’s syndrome, and POLG mutations.
MELAS (mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes) is a rare syndrome caused by 17 different mutations in mitochondrial DNA.6 While no two patients have exactly the same clinical presentation, symptoms can include muscle weakness, strokes, headaches, and anorexia.
Leigh’s syndrome, in contrast, results from mutations in over 30 different nuclear and mitochondrial genes.7 Children afflicted with this syndrome may show no signs in the early stages of their development. However, after a traumatic insult, such as illness or infection, these children develop lesions in their central nervous systems, followed by a decline in cognitive development. Currently, there are no effective therapies for this syndrome, which is in most cases fatal.
The final class of diseases discussed results from a mutation in the nuclear gene POLG, which encodes the mitochondrial DNA polymerase-γ, an enzyme necessary for synthesis of mitochondrial DNA. Disruption of this gene is associated with multiple disorders, including Alper-Huttenlocher Syndrome and myoclonic epilepsy myopathy sensory ataxia. Symptoms range from seizures and psychomotor impairments to blindness.8
Dr. Krotoski concluded her talk by elaborating on measures that NICHD researchers are taking to ease the difficulties of identifying and diagnosing mitochondrial disease. The NICHD is involved in the Mitochondrial Disease Sequence Data Repository Consortium (MseqDR), a collaboration of mitochondrial biologists from around the world who collect and make available phenotypic and genotypic data of various mitochondrial diseases.9,10 NICHD also provides support to the North American Mitochondrial Disease Consortium (NAMDC), a part of the NIH Rare Diseases Consortium. The group includes a network of clinical investigators and clinicians studying mitochondrial disease, patient registries, and clinical trials related to mitochondrial diseases.11
Dr. Neelakanta (Ravi) Ravindranath concluded the NICHD Exchange with his talk entitled “Mitochondrial DNA replacement for the treatment of mitochondrial diseases and infertility.” He presented the potential of restoring fertility to individuals suffering from mitochondrial disease using a technique called spindle transfer.
Mitochondria, and the DNA contained within, are inherited exclusively from an individual’s mother. The egg contains approximately 100,000 of these organelles, all of which are maternally derived; conversely, sperm contribute (almost) no mitochondria to newly fertilized eggs. The mitochondrial health of the mother is therefore a crucial component in determining the health and, in some cases, viability of her children.12
Spindle transfer offers promise in restoring fertility to patients suffering from mitochondrial disease, specifically from mutations in the mitochondrial DNA. This procedure involves replacement of the nucleus in oocytes from healthy donors with nuclear DNA from a patient’s own oocytes prior to in vitro fertilization. This creates healthy oocytes containing the patient’s nuclear DNA and the donor’s healthy mitochondria. While researchers have used this approach to generate viable and healthy transmitochondrial monkeys,13 it has only been used with human oocytes to generate blastocysts and embryonic stem cell lines (in non-federally funded studies).14
Spindle transfer, while promising, is not free from either technical or ethical issues. One limitation of this technique is that small amounts of diseased mitochondria can be carried over with the nuclear DNA, which can result in low levels of heteroplasmy—a situation that arises when cells contain mixed populations of healthy and mutant mitochondrial DNA.15 Their proportion in the cell relative to their healthy counterparts, however, would be low.
A major ethical issue with spindle transfer is that this technique would result in the generation of offspring having DNA from three genetic sources: the mother, the father, and the donor. The effects that this would have on the development and characteristics of a future individual are unknown.
Dr. Ravindranath concluded his talk with the clinical status of cytoplasmic transfer in the United States and United Kingdom. While cytoplasmic transfer is not currently permitted in the United States as a means of assisted reproductive technology, the FDA is currently considering it. A committee of the Institute of Medicine has been appointed to discuss the social and ethical issues of the procedure. On February 3, 2015 (one day after this NICHD Exchange was held), the House of Commons voted to approve spindle transfer for use in the United Kingdom, however it has yet to be approved in the House of Lords.
The role played by mitochondria in the health of both the cell and of the entire individual is complex and multifaceted, yet it is part of a very delicate system in which the smallest of perturbations can have drastic consequences. While mitochondrial disease is mercifully rare, it does cause great hardship for the individuals who suffer from it; yet the difficulty in diagnosing mitochondrial disease may suggest it is not as rare as currently thought.
The collaboration of many minds, and the fusion of multiple perspectives, will be required to combat mitochondrial disease and give those suffering from it better prospects for survival and quality of life. And thus ensure that NICHD fulfills its mission to ensure that every person is born healthy and wanted, that women suffer no harmful effects from reproductive processes, and that all children have the chance to achieve their full potential for healthy and productive lives, free from disease or disability.
- Alberts B, et al. Molecular biology of the Cell. 4th edition. Garland Science NY 2002.
- Zimorski V, et al. Endosymbiotic theory for organelle origins. Curr Opin Microbiol. 2014 Dec;22C:38-48.
- Maio N, et al. Cochaperone binding to LYR motifs confers specificity of iron sulfur cluster delivery. Cell Metab. 2014.19(3):445-457.
- Mochel F, et al. Splice mutation in the iron-sulfur cluster scaffold protein ISCU causes myopathy with exercise intolerance. Am J Hum Genet. 2008 2(3):652-60.
- Rambold AS et al. Tubular network formation protects mitochondria from autophagosomal degradation during nutrient starvation. Proc Natl Acad Sci U S A. 2011 108(25):10190-5.
- National Library of Medicine (US). Genetics Home Reference [Internet]. Bethesda (MD): The Library; 2015 Feb 03. Mitochondrial Encephalomyopathy, Lactic Acidosis, and Stroke Like Episodes; [reviewed 2013 Dec; cited 2014 Feb 15]; [about 4 screens]. Available from: http://ghr.nlm.nih.gov/condition/mitochondrial-encephalomyopathy-lactic-acidosis-and-stroke-like-episodes
- National Library of Medicine (US). Genetics Home Reference [Internet]. Bethesda (MD): The Library; 2015 Feb 09. Leigh Syndrome; [reviewed 2011 Oct; cited 2014 Feb 15]; [about 4 screens]. Available from: http://ghr.nlm.nih.gov/condition/leigh-syndrome
- National Library of Medicine (US). Genetics Home Reference [Internet]. Bethesda (MD): The Library; 2015 Feb 09. POLG; [reviewed 2011 June; cited 2014 Feb 15]; [about 4 screens]. Available from: http://ghr.nlm.nih.gov/gene/POLG
- Falk MJ, et al. Mitochondrial Disease Sequence Data Resource (MSeqDR): A global grass-roots consortium to facilitate deposition, curation, annotation, and integrated analysis of genomic data for the mitochondrial disease clinical and research communities. Mol Genet Metab. 2014. S1096-7192(14)00377-1.
- MseqDR Consortium Overview. Available from: https://mseqdr.org
- North American Mitochondrial Disease Consortium. “Information for Patients and Families.” Available from: http://www.rarediseasesnetwork.org/namdc/
- Shoubridge EA and Wai T. Mitochondrial DNA and the mammalian oocyte. Curr Top Dev Biol. 2007. 77:87-111.
- Tachibana M, et al. Mitochondrial gene replacement in primate offspring and embryonic stem cells. Nature. 2009. 461(7262):367-724.
- Tachibana M, et al. Towards germline gene therapy of inherited mitochondrial diseases. Nature. 2013. 493(7434):627-31.
- Wallace DC and Chalkia D. Mitochondrial DNA genetics and the heteroplasmy conundrum in evolution and disease. Cold Spring Harb Perspect Med. 2013 Oct;3(10):a021220.