Mass spectrometry data for: A small protein coded within the mitochondrial canonical gene nd4 regulates mitochondrial bioenergetics

Background: Mitochondria have a central role in cellular functions, aging and in certain diseases. They possess their own genome, a vestige of their bacterial ancestor. Over the course of evolution, most of the genes of the ancestor have been lost or transferred to the nucleus. In humans, the mtDNA is a very small circular molecule with a functional repertoire limited to only 37 genes. Its extremely compact nature with genes arranged one after the other and separated by short non-coding regions suggests that there is little room for evolutionary novelties. This is radically different from bacterial genomes, which are also circular but much larger, and in which we can find genes inside other genes. These sequences, different from the reference coding sequences, are called alternative open reading frames or altORFs, and they are involved in key biological functions. However, whether altORFs exist in mitochondrial protein-coding genes or elsewhere in the human mitogenome has not been fully..., HeLa cells (n=3 replicates) were washed with PBS and harvested in a lysing buffer containing 10 mM HEPES, 150 mM NaCl and a cocktail of protease inhibitors (benzamidine, PMSF, aprotinin and leupeptin). Cells were then lysed with a sonic dismembrator sonicator (Fisher) 3 times for 30 seconds before adding 100 µl of Triton X-100 10%. Cell lysates were incubated on ice for 20 minutes and then centrifuged for 15 minutes at 15000 RPM at 4˚C. The supernatant was kept. 20 µl of protein A agarose was added to 1 ml of samples and incubated at 4˚C for 1 hour. Samples were then centrifuged at 15000 rpm for 1 minute at 4˚C and the supernatant was kept. 20 µl of rabbit anti-MTALTND4 antibody and 20 µl of protein A agarose were added to the samples before incubating at 4˚C overnight. 20 µl of rabbit pre-immune serum was used as control. Samples were washed 6 times with lysing buffer. Samples were then sent to the proteomics platform of the McGill University Health Center (MUHC) Research Institute for..., ,