RsmD, a Chloroplast rRNA m2G Methyltransferase, Plays a Role in Cold Stress Tolerance by Possibly Affecting Chloroplast Translation in Arabidopsis

Abstract

Ribosomal RNA (rRNA) methylation is a pivotal process in the assembly and activity of ribosomes, which in turn play vital roles in the growth, development and stress responses of plants. Although few methyltransferases responsible for rRNA methylation have been identified in plant chloroplasts, the nature and function of these enzymes in chloroplasts remain largely unknown. In this study, we characterized ArabidopsisRsmD (At3g28460), an ortholog of the methyltransferase responsible for N2-methylguanosine (m2G) modification of 16S rRNA in Escherichia coli. Confocal microscopic analysis of an RsmD- green fluorescent protein fusion protein revealed that RsmD is localized to chloroplasts. Primer extension analysis indicated that RsmD is responsible for m2G methylation at position 915 in the 16S rRNA of Arabidopsis chloroplasts. Under cold stress, rsmd mutant plants exhibited retarded growth, i.e. had shorter roots, lower fresh weight and pale-green leaves, compared with wild-type (WT) plants. However, these phenotypes were not detected in response to drought or salt stress. Notably, the rsmd mutant was hypersensitive to erythromycin or lincomycin and accumulated fewer chloroplast proteins compared with the WT, suggesting that RsmD influences translation in chloroplasts. Complementation lines expressing RsmD in the rsmd mutant background recovered WT phenotypes. Importantly, RsmD harbored RNA methyltransferase activity. Collectively, the findings of this study indicate that RsmD is a chloroplast 16S rRNA methyltransferase responsible for m2G915 modification that plays a role in the adaptation of Arabidopsisto cold stress.

Keywords: Abiotic stress •; Arabidopsis thaliana •; Chloroplast •; RNA modification •; RsmD; rRNA methylation •.