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CSP41a, a multifunctional RNA‐binding protein, initiates mRNA turnover in tobacco chloroplasts
Author(s) -
Bollenbach Thomas J.,
Tatman Dana A.,
Stern David B.
Publication year - 2003
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1046/j.1365-313x.2003.01935.x
Subject(s) - antisense rna , endoribonuclease , rna , biology , messenger rna , microbiology and biotechnology , transcription (linguistics) , nicotiana tabacum , chloroplast , wild type , rna binding protein , biochemistry , mutant , rnase p , gene , linguistics , philosophy
Summary Expression of c hlorplast s tem‐loop binding p rotein (CSP)41a, a highly conserved chloroplast endoribonuclease, was reduced >90% by the expression of antisense RNA in Nicotiana tabacum . The most striking effects of this silencing were two‐ to sevenfold decreases in the degradation rates of rbc L, psb A, and pet D transcripts in lysed chloroplast extracts. These results are consistent with the hypothesis that CSP41a participates in initiating mRNA turnover through endonucleolytic cleavages. Surprisingly, rbc L and psb A mRNAs accumulated to similar levels in wild‐type and antisense lines. This suggested that decreased degradation was compensated by reduced transcription, which was confirmed using run‐on transcription assays. The collective accumulation of pet D‐containing mRNAs in antisense plants decreased by 25% compared to wild‐type controls. However, the relative levels of pet D processing intermediates in wild‐type and antisense plants did not differ, and there were no changes in pet D 3′‐end maturation, suggesting that CSP41a is not required for pet D RNA processing. CSP41a is a Mg 2+ ‐dependent enzyme; therefore, extracts from antisense plants were tested at different Mg 2+ concentrations. These experiments showed that the half‐life of rbc L decreased as the Mg 2+ concentration was reduced, and at <1 m m free Mg 2+ , conditions where CSP41a is nearly inactive in vitro , the rbc L degradation rate was similar in wild‐type and antisense extracts, suggesting that CSP41a is normally bypassed under these conditions. Mg 2+ has been shown to mediate RNA stability during chloroplast biogenesis, and our data suggest that regulation of CSP41a activity by Mg 2+ is a component of this process.