POLYADENYLATION ACCELERATES DEGRADATION OF CHLOROPLAST MESSENGER-RNA

The expression of chloroplast genes is regulated by several mechanisms , one of which is the modulation of RNA stability. To understand how t his regulatory step is controlled during chloroplast development, we h ave begun to define the mechanism of plastid mRNA degradation. We show here that the degradation petD mRNA involves endonucleolytic cleavage at specific sites upstream of the 3' stem-loop structure. The endonuc leolytic petD cleavage products can be polyadenylated in vitro, and si milar polyadenylated RNA products are detectable in vivo. PCR analysis of the psbA and psaA-psaB-rps14 operons revealed other polyadenylated endonucleolytic cleavage products, indicating that poly(A) addition a ppears to be an integral modification during chloroplast mRNA degradat ion. Polyadenylation promotes efficient degradation of the cleaved pet D RNAs by a 3'-5' exoribonuclease. Furthermore, polyadenylation also p lays an important role in the degradation of the petD mRNA 3' end. Alt hough the 3' end stem-loop is usually resistant to nucleases, adenylat ion renders the secondary structure susceptible to the 3'-5' exoribonu clease. Analysis of 3' ends confirms that polyadenylation occurs in vi vo, and reveals that the extent of adenylation increases during the de gradation of plastid mRNA in the dark. Based on these results, we prop ose a novel mechanism for polyadenylation in the regulation of plastid mRNA degradation.