Abstract
RNase P RNA (RPR), the catalytic subunit of the essential RNase P ribonucleoprotein, removes the 5′ leader from precursor tRNAs. The ancestral eukaryotic RPR is a Pol III transcript generated with mature termini. In the branch of the arthropod lineage that led to the insects and crustaceans, however, a new allele arose in which RPR is embedded in an intron of a Pol II transcript and requires processing from intron sequences for maturation. We demonstrate here that the Drosophila intronic-RPR precursor is trimmed to the mature form by the ubiquitous nuclease Rat1/Xrn2 (5′) and the RNA exosome (3′). Processing is regulated by a subset of RNase P proteins (Rpps) that protects the nascent RPR from degradation, the typical fate of excised introns. Our results indicate that the biogenesis of RPR in vivo entails interaction of Rpps with the nascent RNA to form the RNase P holoenzyme and suggests that a new pathway arose in arthropods by coopting ancient mechanisms common to processing of other noncoding RNAs.
Highlights
RNase P is an essential endoribonuclease that is required to cleave the 5 leader of precursor tRNAs [1–3]
To study if RNase P RNA (RPR) maturation is influenced by splicing, we assayed the effects of either depletion of lariat debranching enzyme (Ldbr) or alteration of splice sites
We expressed a split-Red Fluorescent Protein (RFP) reporter gene with the D. virilis intron-encoding RPR inserted between two RFP exons (Dv RPR; Figure 1A)
Summary
RNase P is an essential endoribonuclease that is required to cleave the 5 leader of precursor tRNAs [1–3]. The ribonucleoprotein (RNP) form of RNase P is widespread in all three domains of life [1–3]. The RNP consists of a catalytic RNA (RNase P RNA, RPR) and a variable number of protein subunits (RNase P proteins, Rpps). The number of Rpps associated with the RPR increases from one in bacteria, to four or five in archaea, and eight to ten in eukaryotes [3–5]. The additional Rpps appear to compensate for structural features present only in bacterial RPRs [2,3]. Biochemical reconstitution [6–8] and high-resolution structural [9–11] studies of RNase P from all three domains of life shed light on protein-aided RNA catalysis in this ancient RNP enzyme. Little is known about the sequence of assembly events in vivo for the multi-subunit eukaryotic RNase P holoenzyme. We report the discovery of a critical role for select Rpps in Drosophila RPR biogenesis
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