Deficiency of mitoribosomal S10 protein affects translation and splicing in Arabidopsis mitochondria

Abstract

The ribosome is not only a protein-making machine, but also a regulatory element in protein synthesis. This view is supported by our earlier data showing that Arabidopsis mitoribosomes altered due to the silencing of the nuclear RPS10 gene encoding mitochondrial ribosomal protein S10 differentially translate mitochondrial transcripts compared with the wild-type. Here, we used ribosome profiling to determine the contribution of transcriptional and translational control in the regulation of protein synthesis in rps10 mitochondria compared with the wild-type ones. Oxidative phosphorylation system proteins are preferentially synthesized in wild-type mitochondria but this feature is lost in the mutant. The rps10 mitoribosomes show slightly reduced translation efficiency of most respiration-related proteins and at the same time markedly more efficiently synthesize ribosomal proteins and MatR and TatC proteins. The mitoribosomes deficient in S10 protein protect shorter transcript fragments which exhibit a weaker 3-nt periodicity compared with the wild-type. The decrease in the triplet periodicity is particularly drastic for genes containing introns. Notably, splicing is considerably less effective in the mutant, indicating an unexpected link between the deficiency of S10 and mitochondrial splicing. Thus, a shortage of the mitoribosomal S10 protein has wide-ranging consequences on mitochondrial gene expression.

An Update on Mitochondrial Ribosome Biology: The Plant Mitoribosome in the Spotlight

“…The development of mitoribosome profiling, an experimental approach using deep sequencing of mitochondrial ribosome-protected fragments (RPFs), has allowed revealing aspects of the mitoribosome interactions with the mtmRNA. In yeast, mitochondrial RPFs were around 38 nt long [53], while a bimodal length distribution of RPFs was found in the human [54], maize [55] and Arabidopsis [50]. Data for human mitoribosomes showed a major peak at 27 nt and a minor peak at 33 nt while the maize RPFs showed peaks at 28-29 nt and 36 nt.…”

“…A mitoribosomal profiling analysis of the Arabidopsis translatome has revealed strikingly different mitoribosome association levels with different mtRNAs. mRNAs encoding subunits of OXPHOS complexes showed much higher ribosome loading levels compared to mitochondrial transcripts of ribosomal proteins or those encoding c-type cytochrome maturation factors, implying that the OXPHOS proteins are translated preferentially [50,56]. Low levels of mitoribosome footprints were detected for maturase (MatR) and the TatC protein also known as MttB although these proteins are not identified by mass spectrometry [57].…”

“…In papers from our laboratory [46,49,50] we have shown that the alteration in mitoribosomal protein composition caused by gene silencing can affect the functioning of the mitoribosome. The altered mitoribosomes are able to perform their basic activities (produce proteins), but they differ functionally from the wild-type ones and favor a specific translational program.…”

“…Data for human mitoribosomes showed a major peak at 27 nt and a minor peak at 33 nt while the maize RPFs showed peaks at 28-29 nt and 36 nt. Recent ribosome profiling for Arabidopsis indicates that the majority of mitochondrial ribosome footprints measure 28 nt while reads of 31-32 nt are much less frequent [50]. Moreover, it was shown that the Arabidopsis mitoribosomes altered due to the S10 protein deficiency protect shorter mRNA fragments, suggesting that the size of the RPFs reflects distinct conformations of the mitoribosomes [50].…”

“…Low levels of mitoribosome footprints were detected for maturase (MatR) and the TatC protein also known as MttB although these proteins are not identified by mass spectrometry [57]. Interestingly, the altered mitoribosomes in rps10, markedly more efficiently synthesized the MatR and TatC proteins [50].…”