RESUMO
Modified nucleotides in tRNAs are important determinants of folding, structure and function. Here we identify METTL8 as a mitochondrial matrix protein and active RNA methyltransferase responsible for installing m3C32 in the human mitochondrial (mt-)tRNAThr and mt-tRNASer(UCN). METTL8 crosslinks to the anticodon stem loop (ASL) of many mt-tRNAs in cells, raising the question of how methylation target specificity is achieved. Dissection of mt-tRNA recognition elements revealed U34G35 and t6A37/(ms2)i6A37, present concomitantly only in the ASLs of the two substrate mt-tRNAs, as key determinants for METTL8-mediated methylation of C32. Several lines of evidence demonstrate the influence of U34, G35, and the m3C32 and t6A37/(ms2)i6A37 modifications in mt-tRNAThr/Ser(UCN) on the structure of these mt-tRNAs. Although mt-tRNAThr/Ser(UCN) lacking METTL8-mediated m3C32 are efficiently aminoacylated and associate with mitochondrial ribosomes, mitochondrial translation is mildly impaired by lack of METTL8. Together these results define the cellular targets of METTL8 and shed new light on the role of m3C32 within mt-tRNAs.
Assuntos
Anticódon/química , Metiltransferases/genética , Mitocôndrias/genética , RNA Mitocondrial/química , RNA de Transferência de Serina/química , RNA de Transferência de Treonina/química , Anticódon/metabolismo , Pareamento de Bases , Citosina/metabolismo , Regulação da Expressão Gênica , Células HEK293 , Humanos , Metilação , Metiltransferases/metabolismo , Mitocôndrias/metabolismo , Conformação de Ácido Nucleico , Ligação Proteica , Biossíntese de Proteínas , RNA Mitocondrial/genética , RNA Mitocondrial/metabolismo , RNA de Transferência de Serina/genética , RNA de Transferência de Serina/metabolismo , RNA de Transferência de Treonina/genética , RNA de Transferência de Treonina/metabolismo , Transdução de SinaisRESUMO
High-yielding and stress-resistant crops are essential to ensure future food supply. Barley is an important crop to feed livestock and to produce malt, but the annual yield is threatened by pathogen infections. Pathogens can trigger an altered sugar partitioning in the host plant, which possibly leads to an advantage for the pathogen. Hampering these processes represents a promising strategy to potentially increase resistance. We analysed the response of the barley monosaccharide transporter HvSTP13 towards biotic stress and its potential use for plant protection. The expression of HvSTP13 increased on bacterial and fungal pathogen-associated molecular pattern (PAMP) application, suggesting a PAMP-triggered signalling that converged on the transcriptional induction of the gene. Promoter studies indicate a region that is probably targeted by transcription factors downstream of PAMP-triggered immunity pathways. We confirmed that the nonfunctional HvSTP13GR variant confers resistance against an economically relevant biotrophic rust fungus in barley. Our experimental setup provides basal prerequisites to further decode the role of HvSTP13 in response to biological stress. Moreover, in line with other studies, our experiments indicate that the alteration of sugar partitioning pathways, in a host-pathogen interaction, is a promising approach to achieve broad and durable resistance in plants.
