Differential targeting of VDAC3 mRNA isoforms influences mitochondria morphology

Abstract

Intracellular targeting of mRNAs has recently emerged as a prevalent mechanism to control protein localization. For mitochondria, a cotranslational model of protein import is now proposed in parallel to the conventional posttranslational model, and mitochondrial targeting of mRNAs has been demonstrated in various organisms. Voltage-dependent anion channels (VDACs) are the most abundant proteins in the outer mitochondrial membrane and the major transport pathway for numerous metabolites. Four nucleus-encoded VDACs have been identified in Arabidopsis thaliana. Alternative cleavage and polyadenylation generate two VDAC3 mRNA isoforms differing by their 3′ UTR. By using quantitative RT-PCR and in vivo mRNA visualization approaches, the two mRNA variants were shown differentially associated with mitochondria. The longest mRNA presents a 3′ extension named alternative UTR (aUTR) that is necessary and sufficient to target VDAC3 mRNA to the mitochondrial surface. Moreover, aUTR is sufficient for the mitochondrial targeting of a reporter transcript, and can be used as a tool to target an unrelated mRNA to the mitochondrial surface. Finally, VDAC3-aUTR mRNA variant impacts mitochondria morphology and size, demonstrating the role of mRNA targeting in mitochondria biogenesis.mRNA localization | mRNA sorting | green RNA | porin | plant E ukaryotic cells are characterized by compartmentalization of functions in complex organelles, and cell activity depends on synthesis and precise localization of proteins. The mechanisms of protein sorting are the subject of intense research, and models are often reevaluated (1). The earliest models for protein localization were based on the presence of targeting signals in the polypeptide sequences. However, the targeting of mRNAs to various subcellular regions has recently emerged as an essential regulatory step for protein localization (2).Mitochondria contain hundreds of proteins, but only a few are encoded by the mitochondrial genome. The other proteins are nucleus-encoded and imported into the organelle. Protein import into mitochondria is controlled by peptide sequences that are recognized by mitochondrial receptors (3, 4). In parallel with this posttranslational model, a cotranslational model is now emerging (2, 5). A microarray analysis in Saccharomyces cerevisiae has shown that approximately half of the mRNAs encoding mitochondrial proteins are found in the vicinity of mitochondria (6). Mitochondrial targeting of mRNA was also demonstrated in mammals and plants (7,8).Mechanisms of mitochondrial targeting of mRNAs were essentially studied in yeast. In this species, three factors have been clearly identified to influence mRNA targeting to the mitochondrial surface. (i) Targeting of many but not all mRNAs appears dependent on translation (9, 10). (ii) The RNA-binding protein Puf3p is essential for mitochondrial targeting of a subset of mRNAs (9). (iii) The translocase of the outer mitochondrial membrane (TOM complex) is also involved in mRNA binding (10-12). In addition, some mRNAs are...

Stable nuclear expression ofATP8andATP6genes rescues a mtDNA Complex Vnullmutant

“…Indeed, our group has thus far failed, after many attempts, to express and import CYB into the mitochondria (data not shown). We hope, however, that with careful experimentation and additional levels of targeting (such as RNA targeting to promote co-translational import) (5,62), that this approach will someday become generalized to any of the 13 proteins of the mitochondrial genome.…”

Localized translation near the mitochondrial outer membrane: An update

“…79 More recently, cellular fractionations and in vivo imaging using the MS2-tagging system revealed that VDAC3 mRNAs are associated with mitochondria in Arabidopsis thaliana. 80 Interestingly, VDAC3 transcripts that varied in their 3 0 -UTR length differed in their association, and a region of 140 nts from the 3 0 -UTR appeared sufficient to induce localization of a heterologous mRNA to the mitochondria. These results indicate that localization of VDAC3 mRNA can occur in a non-translational manner, via cis elements from the 3 0 -UTR.…”

“…30,44 Studies from numerous taxa have demonstrated the importance of localized translation to mitochondrial activity. In Arabadopsis thaliana, interference with the localization of mRNA of the membrane protein VDAC3 affected mitochondrial morphology and size 80 and in yeast, the correct localization of ATP2 mRNA was shown to be important for respiration. 41 In human fibroblasts, induced localization of ATP6 mRNA to the mitochondrial outer membrane resulted in the rescue of mitochondrial defects.…”

Early stages in the biogenesis of eukaryotic β‐barrel proteins

“…In yeast, the mRNAs of Tom40, Tob55 and the two isoforms of Porin showed little or no mitochondrial localization, while the mRNA of another β‐barrel protein, mitochondrial distribution and morphology 10 (Mdm10) seemed to be partially enriched in the vicinity of mitochondria . In plant cells, mRNA encoding the β‐barrel protein VDAC3 was shown to be associated with mitochondria . However, while a cotranslational import leads to an mRNA localization to the site of import, the reverse is not necessarily true.…”

“…However, while a cotranslational import leads to an mRNA localization to the site of import, the reverse is not necessarily true. In fact, targeting of VDAC3 mRNA to mitochondria is not required for an efficient import of the protein .…”