Chloroplast RNase J compensates for inefficient transcription termination by removal of antisense RNA

Abstract

Ribonuclease J is an essential enzyme, and the Bacillus subtilis ortholog possesses both endoribonuclease and 59 / 39 exoribonuclease activities. Chloroplasts also contain RNase J, which has been postulated to participate, as both an exo-and endonuclease, in the maturation of polycistronic mRNAs. Here we have examined recombinant Arabidopsis RNase J and found both 59 / 39 exoribonuclease and endonucleolytic activities. Virus-induced gene silencing was used to reduce RNase J expression in Arabidopsis and Nicotiana benthamiana, leading to chlorosis but surprisingly few disruptions in the cleavage of polycistronic rRNA and mRNA precursors. In contrast, antisense RNAs accumulated massively, suggesting that the failure of chloroplast RNA polymerase to terminate effectively leads to extensive symmetric transcription products that are normally eliminated by RNase J. Mung bean nuclease digestion and polysome analysis revealed that this antisense RNA forms duplexes with sense strand transcripts and prevents their translation. We conclude that a major role of chloroplast RNase J is RNA surveillance to prevent overaccumulation of antisense RNA, which would otherwise exert deleterious effects on chloroplast gene expression.

The Primary Transcriptome of Barley Chloroplasts: Numerous Noncoding RNAs and the Dominating Role of the Plastid-Encoded RNA Polymerase

“…Moreover, asRNAs generated as byproducts of read-through transcription and subsequent processing can interfere with the translational efficiency of the target mRNAs. Such asRNAs are normally eliminated by RNase J (Sharwood et al, 2011).…”

The Conserved Endoribonuclease YbeY Is Required for Chloroplast Ribosomal RNA Processing in Arabidopsis

“…For example, the plant-specific endoribonucleases Chloroplast Stem-Loop Binding Protein 41a (CSP41a) and CSP41b have been reported to interact with each other and to be associated with ribosomes (Beligni and Mayfield, 2008;Bollenbach et al, 2009). Chloroplast RNase J, which displays both endoribonucleolytic and exoribonucleolytic activities, plays a major role in RNA surveillance and 59 mRNA maturation (Sharwood et al, 2011;Luro et al, 2013). In addition, RNase J deficiency results in accumulation of 16S and 23S rRNAs that carry 59 extensions (Sharwood et al, 2011).…”

SCO5745, a Bifunctional RNase J Ortholog, Affects Antibiotic Production in Streptomyces coelicolor

“…RNases J have also been characterized in the Gram-negative bacterium Sinorhizobium (7), in the hyperthermophilic archaea Pyrococcus abyssi and Thermococcus kodakaraensis (8), and in plant chloroplasts (9). B. subtilis expresses two paralogous proteins, RNase J1 and J2.…”

“…However, the two proteins form a heterotetramer in vivo that cleaves a model substrate at a different site than the J1 homodimer (10). The single-stranded exonuclease activity of RNases J appears to be strongly dependent upon the phosphorylation state of the 5= end of transcripts, usually showing a distinct preference for the monophosphorylated state (4,9,11,12).…”

“…It has been shown in several systems that RNases J are active in rRNA maturation (5,7). Chloroplast RNase J functions as an RNA surveillance mechanism for antisense RNA (9). In B. subtilis, an RNase J2 null mutation combined with reduced expression of RNase J1 globally affected the expression of over 600 genes (46).…”