Suborganelle localization, interaction, and viability fine-tuning patterns of the human complete set of mitochondrial aminoacyl-tRNA synthetases

On December 12, the international academic journal Nucleic Acids Research published online the latest collaborative research results of Xiaolong Zhou's research group and Wang Enduo's research group of the Center for Excellence in Molecular Cell Science (Institute of Biochemistry and Cell Biology) of the Chinese Academy of Sciences, "RNA granule-clustered mitochondrial aminoacyl-tRNA synthetases form multiple.
" complexes with the potential to fine-tune tRNA aminoacylation”
。

The human genome contains 37 aminoacyl-tRNA synthetase (aaRS) genes, responsible for encoding all aaRSs
used in both cytoplasmic and mitochondrial protein synthesis systems.
aaRS catalyzes the tRNA aminoacylation reaction to generate aminoacyl-tRNA, ensuring the speed and accuracy of
protein synthesis.
In contrast to bacterial-derived aaRS, mammalian cytoplasmic aaRS usually contains N- or C-terminal extensions, which mediate protein-protein interactions, such as 9 human cytoplasmic aaRS and 3 non-enzymatic cofactors to form cytoplasmic multi-aaRS complexes (MSCs).

The formation of MSCs is of great significance for the efficiency of
protein synthesis and the non-protein synthesis function of aaRS.
In addition to human cytoplasmic MSCs, simple aaRS-containing complexes or MSCs
are present in bacteria, archaea and yeast.
Nineteen human mitochondrial aaRSs are encoded
by nuclear genes.
Although the whole set of human mitochondrial aaRS has been identified 20 years ago, the suborganelle localization and interaction protein network of human mitochondrial aaRS has not been systematically established; The mode of activity regulation of human mitochondrial aaRS is also unclear
.

Mitochondrial RNA granules (MRG) are dynamic suborganelle structures composed of proteins and RNA discovered in recent years, which are involved in the processing, maturation and production
of mitochondrial RNA.
Proteomic and fluorescence experiments of MRG reveal multiple mitochondrial protein distributions associated with RNA metabolism, including tRNA processing enzymes RNase P and RNase Z, rRNA methyltransferase, and tRNA modifiers
.
Although multiple mitochondrial aaRSs have been found in MRG proteomics, specific experimental evidence
has been lacking.

In this study, the researchers first analyzed the primary structure of human mitochondrial aaRS and bacterial and eukaryotic cytoplasmic aaRS, revealing that human mitochondrial aaRS is evolutionarily more homologous to bacteria-derived aaRS.
cloning and expression of the full set of mitochondrial aaRS genes; By confocal and super-resolution microscopy, 19 mitochondrial aaRS were partially localized in MRG.
Further construction of mitochondrial aaRS mutants without tRNA binding ability, revealing that tRNA binding ability is essential for mitochondrial aaRS localization in MRG.
The interaction network of 19 human mitochondrial aaRSs was systematically established by immunoprecipitation and mass spectrometry (IP-MS).
Further methods of co-immunoprecipitation, protein co-purification, and in vitro recombination revealed that multiple mitochondrial aaRS forms MSCs, including mitochondrial ala-tRNA synthetase (mtAlaRS)-mitochondrial seryl-tRNA synthetase (mtSerRS) interaction, mitochondrial asparagine-tRNA synthetase (mtAsnRS)-mtSerRS interaction, mitochondrial tyrosyl-tRNA synthetase (mtTyrRS)- mtSerRS interaction, mitochondrial arginyl-tRNA synthetase (mtArgRS)-mitochondrial threonyl-tRNA synthetase (mtThrRS) interaction, etc.
; Furthermore, in vitro viability recombination experiments proved that the formation of mitochondrial MSCs has a regulating effect on the aminoacylation activity of specific subunits
.

In this study, all mitochondrial aaRS were partially localized in MRG, and the potential molecular basis for mitochondrial aaRS to enter MRG was elucidated.
A complete set of mitochondrial aaRS interaction networks were established and several mitochondrial MSCs were discovered; The synergistic regulation of mitochondrial MSC formation on the aminoacylation efficiency of different mitochondrial tRNAs was revealed
.
These results deepen the understanding of the suborganelle localization, interaction network and viability fine-tuning mechanism of mitochondrial aaRS, and provide new insights
into the molecular mechanism of mitochondrial aaRS gene mutations leading to related diseases.

Peng Guixin, a doctoral student jointly trained by the Center of Excellence for Molecular Cells and ShanghaiTech University, and Xueling Mao, a doctoral student at the Center of Excellence in Molecular Cells, are co-first authors of this paper, and researchers Xiaolong Zhou and Enduo Wang are co-corresponding authors
of this paper.
Dr.
Qingrun Li from the Center of Excellence for Molecular Cells and Professor Xin Chen from Xiamen University participated in the study
.
The work was funded by the National Key Research and Development Program of the Ministry of Science and Technology, the Foundation Committee, the Chinese Academy of Sciences, and the Shanghai Municipality
.

Article link: https://academic.
oup.
com/nar/advance-article/doi/10.
1093/nar/gkac1141/6887605

Suborganelle localization, interaction, and viability fine-tuning mechanisms of mitochondrial aaRS