bims-mitran 2022-03-13 papers

Issue of 2022–03–13
three papers selected by
Andreas Kohler, Stockholm University

RNA. 2022 Mar 07. pii: rna.079097.122. [Epub ahead of print]

Mito-FUNCAT-FACS reveals cellular heterogeneity in mitochondrial translation.

Yusuke Kimura, Hironori Saito, Tatsuya Osaki, Yasuhiro Ikegami, Taisei Wakigawa, Yoshiho Ikeuchi, Shintaro Iwasaki.

Mitochondria possess their own genome that encodes components of oxidative phosphorylation (OXPHOS) complexes, and mitochondrial ribosomes within the organelle translate the mRNAs expressed from the mitochondrial genome. Given the differential OXPHOS activity observed in diverse cell types, cell growth conditions, and other circumstances, cellular heterogeneity in mitochondrial translation can be expected. Although individual protein products translated in mitochondria have been monitored, the lack of techniques that address the variation in overall mitochondrial protein synthesis in cell populations poses analytic challenges. Here, we adapted mitochondrial-specific fluorescent noncanonical amino acid tagging (FUNCAT) for use with fluorescence-activated cell sorting (FACS) and developed mito-FUNCAT-FACS. The click chemistry-compatible methionine analog L-homopropargylglycine (HPG) enabled the metabolic labeling of newly synthesized proteins. In the presence of cytosolic translation inhibitors, HPG was selectively incorporated into mitochondrial nascent proteins and conjugated to fluorophores via the click reaction (mito-FUNCAT). The application of in situ mito-FUNCAT to flow cytometry allowed us to separate changes in net mitochondrial translation activity from those of the organelle mass and detect variations in mitochondrial translation in cancer cells. Our approach provides a useful methodology for examining mitochondrial protein synthesis in individual cells.

Keywords: FACS; FUNCAT; HPG; Mitochondria; Translation

DOI: https://doi.org/10.1261/rna.079097.122

Sci Rep. 2022 Mar 08. 12(1): 4056

Mitochondrial dysfunction-induced high hCG associated with development of fetal growth restriction and pre-eclampsia with fetal growth restriction.

Ryo Kiyokoba, Takeshi Uchiumi, Mikako Yagi, Takahiro Toshima, Shigehiro Tsukahara, Yasuyuki Fujita, Kiyoko Kato, Dongchon Kang.

Fetal growth restriction (FGR) and pre-eclampsia with fetal growth restriction (PE/FGR) are high-risk perinatal diseases that may involve high levels of human chorionic gonadotropin (hCG) and mitochondrial dysfunction. However, little is known about how these factors affect placental function. We investigated how mitochondrial dysfunction and high hCG expression affected placental function in unexplained FGR and PE/FGR. We observed elevated expression of hCGβ and growth differentiation factor 15 mRNA and protein levels in the placenta with both diseases. Likewise, antiangiogenic factors, such as Ang2, IP10, sFlt1, IL8, IL1B, and TNFα, were also upregulated at the mRNA level. In addition, the expression of COXI and COXII which encoded by mitochondrial DNA were significantly decreased in both diseases, suggesting that mitochondrial translation was impaired. Treatment with hCG increased Ang2, IP10, IL8, and TNFα mRNA levels in a dose-dependent manner via the p38 and JNK pathways. Mitochondrial translation inhibitors increased hCGβ expression through stabilization of HIF1α, and increased IL8 and TNFα mRNA expression. These results revealed that high expression of hCG due to mitochondrial translational dysfunction plays an important role in the pathogenesis of FGR and PE/FGR.

DOI: https://doi.org/10.1038/s41598-022-07893-y

Nat Commun. 2022 Mar 11. 13(1): 1275

Functional genomics of RAP proteins and their role in mitoribosome regulation in Plasmodium falciparum.

Thomas Hollin, Steven Abel, Alejandra Falla, Charisse Flerida A Pasaje, Anil Bhatia, Manhoi Hur, Jay S Kirkwood, Anita Saraf, Jacques Prudhomme, Amancio De Souza, Laurence Florens, Jacquin C Niles, Karine G Le Roch.

The RAP (RNA-binding domain abundant in Apicomplexans) protein family has been identified in various organisms. Despite expansion of this protein family in apicomplexan parasites, their main biological functions remain unknown. In this study, we use inducible knockdown studies in the human malaria parasite, Plasmodium falciparum, to show that two RAP proteins, PF3D7_0105200 (PfRAP01) and PF3D7_1470600 (PfRAP21), are essential for parasite survival and localize to the mitochondrion. Using transcriptomics, metabolomics, and proteomics profiling experiments, we further demonstrate that these RAP proteins are involved in mitochondrial RNA metabolism. Using high-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (eCLIP-seq), we validate that PfRAP01 and PfRAP21 are true RNA-binding proteins and interact specifically with mitochondrial rRNAs. Finally, mitochondrial enrichment experiments followed by deep sequencing of small RNAs demonstrate that PfRAP21 controls mitochondrial rRNA expression. Collectively, our results establish the role of these RAP proteins in mitoribosome activity and contribute to further understanding this protein family in malaria parasites.

DOI: https://doi.org/10.1038/s41467-022-28981-7