Small RNAs reveal two target sites of the RNA-maturation factor Mbb1 in the chloroplast

Many chloroplast transcripts are protected against exonucleolytic degradation by RNA-binding proteins. Such interactions can lead to the accumu-lation of short RNAs (sRNAs) that represent foot-prints of the protein partner. By mining existing data sets of Chlamydomonas reinhardtii small RNAs, we identify chloroplast sRNAs. Two of these correspond to the 50-ends of the mature psbB and psbH messenger RNAs (mRNAs), which are both stabilized by the nucleus-encoded protein Mbb1, a member of the tetratricopeptide repeat family. Accordingly, we find that the two sRNAs are absent from the mbb1 mutant. Using chloroplast transformation and site-directed mutagenesis to survey the psbB 50 UTR, we identify a cis-acting element that is essential for mRNA accumulation. This sequence is also found in the 50 UTR of psbH, where it plays a role in RNA processing. The two sRNAs are centered on these cis-acting elements. Furthermore, RNA binding assays in vitro show that Mbb1 associates with the two elements specifically. Taken together, our data identify a conserved cis-acting element at the extremity of the psbH and psbB 50 UTRs that plays a role in the processing and stability of the respective mRNAs through inter-actions with the tetratricopeptide repeat protein Mbb1 and leads to the accumulation of protected sRNAs