Abstract
The initiation of protein synthesis in bacteria is ruled by three canonical factors: IF1, IF2, and IF3. This system persists in human mitochondria; however, it functions in a rather different way due to specialization and adaptation to the organellar micro-environment. We focused on human mitochondrial IF3, which was earlier studied in vitro, but no knock-out cellular models have been published up to date. In this work, we generated human HeLa cell lines deficient in the MTIF3 gene and analyzed their mitochondrial function. Despite the lack of IF3mt in these cells, they preserved functional mitochondria capable of oxygen consumption and protein synthesis; however, the translation of ATP6 mRNA was selectively decreased which compromised the assembly of ATP synthase. Together with the analogous results obtained earlier for baker’s yeast mitochondrial IF3, our findings point to a functional divergence of mitochondrial initiation factors from their bacterial ancestors.
Highlights
Mitochondria are double-membrane, essential organelles of eukaryotic cells responsible for aerobic respiration – a complex chain of redox reactions resulting in the production of ATP, the main cellular energetic currency, which fuels practically all biological processes
In the Online GEene Essentiality database (OGEE, http:// ogee.medgenius.info) we found the information about seven published large-scale CRISPR/Cas[9] and RNAi screenings indicating that the deficiency of the MTIF3 gene should not compromise the survivability and the growth of the human cultured cells
There is a report of the International Mouse Phenotyping Consortium (IMPC) that MTIF3 gene disruption generated in mice had a deleterious phenotype: heterozygous mutants demonstrated hypoactivity and homozygous mutation resulted in the preweaning lethality, and complete penetrance
Summary
Mitochondria are double-membrane, essential organelles of eukaryotic cells responsible for aerobic respiration – a complex chain of redox reactions resulting in the production of ATP, the main cellular energetic currency, which fuels practically all biological processes. Initiation of protein synthesis in bacterial cells is governed by three canonical factors: IF1, IF2, and IF3 These play an essential role in the recruitment of ribosomes on the mRNA, and the correct selection and positioning of the start codon and initiator tRNA5. The www.nature.com/scientificreports activity of the separate C-domain of IF3mt is significantly reduced compared to the full-size protein; at the same time, the N-domain and the linker can bind the 28S subunit[10]. These data indicate that the mechanisms of action of IF3 and IF3mt are different despite their homology and structural similarity. We generated MTIF3 knock-out human cell lines and analyzed their mitochondrial function
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