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A moss pentatricopeptide repeat protein binds to the 3′ end of plastid clpP pre‐mRNA and assists with mRNA maturation
Author(s) -
Hattori Mitsuru,
Sugita Mamoru
Publication year - 2009
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/j.1742-4658.2009.07267.x
Subject(s) - pentatricopeptide repeat , exoribonuclease , biology , messenger rna , rna , rna splicing , intron , rna binding protein , microbiology and biotechnology , rnase p , genetics , gene , rna editing
Pentatricopeptide repeat (PPR) proteins constitute a large family in land plants and are required for various post‐transcriptional steps associated with RNA in plant organelles. The moss Physcomitrella patens PPR protein, PpPPR_38, is a nuclear‐encoded chloroplast protein and was previously shown to be involved in the maturation step of chloroplast clpP pre‐mRNA. To understand precisely the molecular function of PpPPR_38, we prepared recombinant PpPPR_38 protein and characterized it in maturation steps of clpP pre‐mRNA. In vitro RNA‐binding assays showed that the recombinant protein strongly bound to the clpP ‐5′‐ rps12 intergenic region, which is highly AU‐rich and includes an inverted repeat sequence potentially forming a stem‐loop structure. Digestion of the bound RNA region by RNase V1 was significantly accelerated by the addition of the recombinant protein. This strongly suggests that the binding of PpPPR_38 facilitates the formation of a stable stem‐loop structure. An in vitro degradation assay using chloroplast lysates gave rise to the possibility that the stable stem‐loop structure formed by PpPPR_38 contributes the correct intergenic RNA cleavage and protection of mature clpP mRNA against 3′ to 5′ exoribonuclease. Because an RNA‐binding assay also showed weak binding to the clpP first exon–intron region, PpPPR_38 is likely to be related to the splicing of clpP pre‐mRNA. Taking together all of the above findings, we conclude that PpPPR_38 is necessary for several steps in the clpP mRNA maturation process.