Newswise — Imagine trying to figure out how your car's power train works from just a few of its myriad components: It would be nearly impossible. Scientists have long faced a similar challenge in understanding cells' tiny powerhouses — called "mitochondria" — from scant knowledge of their molecular parts.
Now, an international team of researchers has created the most comprehensive "parts list" to date for mitochondria, a compendium that includes nearly 1,100 proteins. By mining this critical resource, the researchers have already gained deep insights into the biological roles and evolutionary histories of several key proteins. In addition, this careful cataloging has identified a mutation in a novel protein-coding gene as the cause behind one devastating mitochondrial disease. A full description of the work appears in the July 11 print edition of the journal Cell.
"For years, a fundamental question in cell biology has gone largely unanswered — what proteins function in mitochondria?" said Vamsi Mootha, an associate member at the Broad Institute of Harvard and MIT and a Harvard Medical School assistant professor at Massachusetts General Hospital, who led the study. "By creating a comprehensive list, we now have a valuable resource that has already helped enhance our understanding of mitochondrial biology and disease."
Mitochondria are linchpins of cellular life, found within the cells of all eukaryotes from yeast to humans. These miniaturized organs ("organelles" ) are well known for their role in providing cellular energy. They have also been implicated in a wide range of normal and disease processes, including diabetes, neurodegeneration, cancer, drug toxicity and aging.
Although mitochondria have their own genome — a vestige from their days as free-living bacteria — the vast majority of the critical mitochondrial proteins are derived not from their genome, but rather from the nuclear genome. However, even with the wealth of genome sequence data now available, scientists have struggled to identify which genes encode the roughly 1,200 proteins that make up a functional mitochondrion.
Researchers from the Broad Institute, Harvard Medical School, and Massachusetts General Hospital worked together to address this problem, drawing on the power of a multi-faceted approach that includes large-scale, mass spectrometry-based proteomics to measure proteins in mitochondria from a variety of tissues; computational methods to help identify those proteins that cannot be reliably detected; and microscopy to confirm within human cells the localization of presumptive mitochondrial proteins.
"The technologies and analytical methods for measuring proteins on a large scale are really transforming what we can learn about human biology," said Steve Carr, director of the Proteomics Platform at the Broad Institute and a co-author of the Cell paper. "By applying them to mitochondria isolated from fourteen different mouse tissues, we've completed one of the most comprehensive proteomic analyses of any organelle to date."
As a result of their analyses, the researchers identified a total of 1,098 mitochondrial proteins to form a compendium they have named "MitoCarta," and which is available to the entire scientific community. Notably, about one-third of this inventory has not been previously linked to the organelle.
To shed light on the functions of the newly uncovered mitochondrial proteins, the researchers compared the proteins' corresponding gene sequences across hundreds of species, from humans and fish to fungi and bacteria. "Proteins with similar roles often share similar histories, meaning they're gained or lost together during evolution," said Mootha. "We decided to use this tendency to our advantage to decipher how some mitochondrial proteins work."
By examining the organelle's proteins through this evolutionary lens, the researchers uncovered a striking pattern. A group of key mitochondrial proteins, known to be absent in yeast but otherwise present among eukaryotes, are actually missing from several other single-celled species. In organisms that have them, including humans and other mammals, these proteins contribute to a boot-shaped, multi-protein structure, which forms the gateway to a critical step in the energy-generation process. By virtue of these proteins' shared — and unusual — past, Mootha and his colleagues were able to identify several additional proteins that are also associated with this crucial mitochondrial structure, known as complex 1.
In addition to offering insights into mitochondrial biology, these discoveries also paved the way for a breakthrough in understanding mitochondrial disease. For decades, doctors have diagnosed patients with deficiencies in complex I function. These disorders affect about 1 in 5,000 newborns, are genetic in origin, and are lethal in the first few years of life. Yet for many cases a culprit gene cannot be found. However, thanks to MitoCarta and its corresponding evolutionary analyses, the researchers and their collaborators at the University of Melbourne and Royal Children's Hospital in Australia identified a mutation in a novel gene, called C8orf38, as one cause of complex I disease.
"Our finding underscores the power of this protein catalogue to open new vistas on disease," said Mootha. "It promises to shed light not only on rare metabolic diseases, but common diseases as well."
This work was supported by a grant from the National Institute of General Medical Sciences, one of the National Institutes of Health.
Data access
The mitochondrial protein compendium (MitoCarta) is freely available to researchers on the web: http://www.broad.mit.edu/publications/MitoCarta. In addition, all of the raw mass spectrometry files are available for download at http://www.proteomecommons.org/data/show.jsp?id=7820.
Paper cited:
Pagliarini DJ et al. A mitochondrial protein compendium elucidates complex I disease biology. Cell July 11, 2008.
About the Broad Institute of MIT and Harvard
The Broad Institute of MIT and Harvard was founded in 2003 to bring the power of genomics to biomedicine. It pursues this mission by empowering creative scientists to construct new and robust tools for genomic medicine, to make them accessible to the global scientific community, and to apply them to the understanding and treatment of disease.
The Institute is a research collaboration that involves faculty, professional staff and students from throughout the MIT and Harvard academic and medical communities. It is jointly governed by the two universities.
Organized around Scientific Programs and Scientific Platforms, the unique structure of the Broad Institute enables scientists to collaborate on transformative projects across many scientific and medical disciplines.
For further information about the Broad Institute, go to http://www.broad.mit.edu.
Media Backgrounder: A Primer on Mitochondria
1. What are mitochondria and where are they found?
Mitochondria are bean-shaped compartments within cells that supply energy. These compartments, a type of membrane-bound organelle, are found in eukaryotes — organisms whose cells have nuclei, the home of the genome. Multicellular organisms (humans, mice, fish, etc.) as well as some unicellular ones, like yeast, are counted as eukaryotes. Bacteria, though, are not: They are considered prokaryotes for their lack of organelles, including mitochondria and nuclei.
Intriguingly, mitochondria vary widely across organisms and even within an organism. Drastic differences can exist in the number of mitochondria per cell, their size and morphology, and even their biochemical capabilities. For example, fatty acids readily broken down by mitochondria in muscle, but not brain tissue. Because of a lack of molecular knowledge about mitochondria and their resident proteins, the basis for such differences is largely unclear.
2. What do mitochondria do?
Although mitochondria are perhaps best known for their roles in energy metabolism, they also participate in a plethora of other key biological processes. These include critical functions such as programmed cell death (or "apoptosis" ), a normal mechanism through which old or damaged cells can be eliminated.
Defects in mitochondria are associated with more than 50 human diseases, ranging from in-born errors of metabolism in infants to neurodegeneration in adults. Moreover, several common diseases, such as cancer and type 2 diabetes, have been associated with mitochondrial dysfunction. Prescription drugs can also disrupt mitochondria. Such drug-induced toxicity is a reason why some drugs are pulled from the market and why some potential drugs fail the clinical trial process.
3. Where do mitochondria come from?
Mitochondria, it turns out, have their own tiny genome. And in humans, this mitochondrial DNA is inherited solely from the mother. Such maternal inheritance arises because mitochondria from sperm are lost following fertilization, while those contributed by the egg persist. Because it is maternally inherited, mitochondrial DNA can provide clues about human history, including the most recent common matrilineal ancestor of living humans (so-called "Mitochondrial Eve" .)
But there are, in fact, paternal contributions to mitochondria. The parts of the mitochondria that are derived from nuclear genes actually come from both parents (see below). This follows a core principle of human genetics: of the 23 pairs of chromosomes that make up your nuclear genome, roughly half come from Mom and the other half from Dad.
Evolutionarily speaking, mitochondria have a very interesting history. They are descendants of an ancient bacterium — a relative of the modern bacterial species, Rickettsia prowazekii — that some 2 billion years ago was enveloped by another cell. That moment marked the beginning of a long and mutually beneficial relationship with eukaryotic cells, known as endosymbiosis. As a result of such "co-habitation" , eukaryotic cells and mitochondria have evolved and adapted to life together, such that now, neither can survive alone.
4. Where do the proteins in mitochondria come from?
Because of the organelle's unusual past, the molecular pieces that make up mitochondria have undergone some shuffling of their own. Mitochondria carry a small circular genome, a vestige of their days as free-living bacteria that has been winnowed during evolution to just a few protein-coding genes.
The human mitochondrial genome was decoded in 1981, a full 20 years before the human genome itself was decoded. The organelle's genome consists of roughly 16,000 chemical units called base pairs, much smaller than the nuclear genome's 3 billion base pairs. The mitochondrial genome includes just 37 genes: 13 genes that encode proteins and 24 additional non-protein coding genes.
The rest of the genes required for a functioning mitochondrion, roughly 1,200 to 1,500 in total, now reside in the nucleus. Identifying these genes from DNA sequence data alone has proven immensely difficult, which is why other large-scale approaches — namely proteomics and computational methods — are required to pinpoint them.
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