When something goes wrong in the mitochondria, the small organelles that feed the cells, it can cause a bewildering variety of symptoms such as poor growth, fatigue and weakness, seizures, cognitive and developmental disabilities, and vision problems. The culprit could be a defect in any of the 1,300 proteins that make up mitochondria, but scientists have very little idea what many of these proteins do, making it difficult to identify the defective protein and treat the disease.
Researchers at the University of Washington School of Medicine in St. Louis and the University of Wisconsin – Madison systematically analyzed dozens of mitochondrial proteins of unknown function and suggested functions for many of them. With these data as a starting point, they identified the genetic causes of three mitochondrial diseases and proposed another 20 possibilities for further research. The findings, published in Nature on May 25, indicate that understanding how hundreds of mitochondrial proteins work together to generate energy and perform other organelle functions could be a promising way to find better ways to diagnose and treat these conditions.
We have a list of parts for mitochondria, but we don’t know what many parts do. It’s like having a problem with your car and taking it to a mechanic, and when you open the hood, they say, “We’ve never seen half of these parts before.” They wouldn’t know how to fix it. This study is an attempt to define the functions of as many of these mitochondrial parts as we can, so that we have a better understanding of what happens when they do not work and, ultimately, a better opportunity to develop therapies to rectify these problems. “
David J. Pagliarini, PhD, co-author, Professor Hugo F. and Ina C. Urbauer, and BJC researcher at the University of Washington
Mitochondrial diseases are a group of rare genetic conditions that collectively affect one in 4,300 people. Because mitochondria provide energy to almost every cell, people with defects in their mitochondria can have symptoms anywhere in the body, although the symptoms are usually more pronounced in tissues that require more energy, such as the heart. the brain and muscles. .
To better understand how mitochondria work, Pagliarini teamed up with colleagues, including lead co-author Joshua J. Coon, PhD, a professor of chemistry and biomolecular chemistry at UW-Madison and a researcher at the Morgridge Institute for Research; and co-authors Jarred W. Rensvold, PhD, former Pagliarini Laboratory Staff Scientist, and Evgenia Shishkova, PhD, Coon Laboratory Staff Scientist, to identify the functions of as many mitochondrial proteins as possible.
The researchers used CRISPR-Cas9 technology to remove individual genes from a human cell line. The procedure created a set of related cell lines, each derived from the same original cell line but with a single gene removed. The missing genes encoded 50 mitochondrial proteins of unknown function and 66 mitochondrial proteins with known functions.
They then examined each cell line to find clues about the role that each missing gene normally plays in keeping the mitochondria functioning properly. The researchers monitored the growth rates of the cells and quantified the levels of 8,433 proteins, 3,563 lipids and 218 metabolites for each cell line. They used the data to create the MITOMICS application (Mitochondrial Orphan Protein Multisome CRISPR Screen), equipping it with tools to analyze and identify biological processes that failed when a specific protein disappeared.
After validating the approach with mitochondrial proteins of known function, the researchers proposed possible biological roles for many mitochondrial proteins of unknown function. With further research, they were able to bind three proteins to three separate mitochondrial conditions.
“It’s very exciting to see how our mass spectrometry technology platform can generate data at this scale, but more importantly, data that can help us directly understand human diseases,” Coon said.
One of the conditions is a multisystem disorder caused by defects in the main path of energy production. Co-author Robert Taylor, PhD, DSc, Professor of Mitochondrial Pathology at the University of Newcastle in Newcastle-upon-Tyne, UK, identified a patient with clear signs of the disorder but no mutations in the usual suspicious genes. The researchers identified a new gene in the pathway and showed that the patient carried a mutation in it.
Separately, Pagliarini and colleagues found that disrupting a gene, RAB5IF, eliminated a protein encoded by a different gene, TMCO1, which has been linked to cerebrophathoracic dysplasia. The condition is characterized by distinctive facial features and severe intellectual disability. In collaboration with co-author Nurten Akarsu, PhD, professor of human genetics at Hacettepe University in Ankara, Turkey, the researchers showed that a mutation in RAB5IF was responsible for one case of cerebrophagiothoracic dysplasia and two cases of cleft lip. in a Turkish family. .
A third gene, when discontinued, caused problems with sugar storage, contributing to a fatal autoinflammatory syndrome. Data on this syndrome were published last year in an article by Bruno Reversade, PhD, of A * STAR, the Singapore Science, Technology and Research Agency.
“We focused mainly on the three conditions, but found data that connected about 20 more proteins to biological pathways or processes,” said Pagliarini, a professor of cell biology and physiology, biochemistry and molecular biophysics, and genetics. “We can’t chase 20 stories in one document, but we made hypotheses and published them so that we and others could prove them.”
To help with scientific discovery, Pagliarini, Coon and colleagues have made the MITOMICS app available to the public. They incorporated several easy-to-use analysis tools so anyone can search for patterns and create plots with just a click. All data can be downloaded for further analysis.
“The hope is that this large data set will become one of the many in the field that will help us collectively come up with better biomarkers and diagnoses for mitochondrial disease,” Pagliarini said. “Every time we discover a function of a new protein, it gives us a new opportunity to orient a pathway therapeutically. Our long-term goal is to understand mitochondria deep enough to be able to intervene therapeutically, which we still can’t do. … “
Source:
University of Washington School of Medicine in St. Louis
Magazine reference:
Rensvold, JW, et al. (2022) Definition of mitochondrial protein functions using a deep multomyomic profile. Nature. doi.org/10.1038/s41586-022-04765-3.