The minor spliceosome is a ribonucleoprotein complex that catalyses the removal of an atypical class of spliceosomal introns (U12-type) from eukaryotic messenger RNAs1,2. It was first identified and characterized in animals, where it was found to contain several unique RNA constituents that share structural similarity with and seem to be functionally analogous to the small nuclear RNAs (snRNAs) contained in the major spliceosome3,4,5,6,7,8. Subsequently, minor spliceosomal components and U12-type introns have been found in plants9,10,11,12 but not in fungi. Unlike that of the major spliceosome, which arose early in the eukaryotic lineage13,14, the evolutionary history of the minor spliceosome is unclear because there is evidence of it in so few organisms. Here we report the identification of homologues of minor-spliceosome-specific proteins and snRNAs, and U12-type introns, in distantly related eukaryotic microbes (protists) and in a fungus (Rhizopus oryzae). Cumulatively, our results indicate that the minor spliceosome had an early origin: several of its characteristic constituents are present in representative organisms from all eukaryotic supergroups for which there is any substantial genome sequence information. In addition, our results reveal marked evolutionary conservation of functionally important sequence elements contained within U12-type introns and snRNAs.
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We thank M. N. Schnare, T. E. Shutt and A. J. Lohan for discussions; M. Dlutek for automated DNA sequencing; and the various genome sequencing centres for making preliminary data publicly available, including the Broad Institute of Harvard and MIT (Rhizopus) and the Joint Genome Institute (Phytophthora). This work was supported by funding (to M.W.G.) from the Canadian Institutes of Health Research and Genome Canada (Protist EST Program). M.W.G. acknowledges salary support from the Canada Research Chairs Program and the Canadian Institute for Advanced Research. Author Contributions J.M.C initially discovered the first protist minor-spliceosomal snRNA (A. castellanii U12) and A.G.R later identified the Phytophthora U11 and U12 snRNAs. A.G.R. did the intron and protein orthologue searches and experimentally verified intron removal and snRNA gene expression. D.F.S. assisted with computer analysis and isolated A. castellanii nucleic acids. M.W.G discovered the COMMD7 intron and initiated a search for minor-spliceosome-specific protein orthologues in protists. A.G.R. wrote the paper. All authors discussed the results and critiqued the final manuscript.
U12-Type Minor spliceosomal introns in Acanthamoeba castellanii. Complete annotation of the sequences, features and gene locations of the U12-type introns identified in the protist A. castellanii
Identification of minor spliceosome-specific protein homologues. Predicted amino acid sequences and alignments of minor spliceosome-specific protein orthologues and sequence-related major spliceosomal proteins identified in this study
U12-Type minor spliceosomal introns in Rhizopus (Fungi) and Phytophthora (Chromalveolata) Complete annotation of the sequences, features, gene locations, and evidence supporting the identification of U12-type introns in P. ramorum, P. sojae, and Rhizopus oryzae
Secondary structures of spliceosomal U2 and U12 snRNAs and detection of Acanthamoeba castellanii U12 snRNA. Sequences and predicted secondary structures of U12 and U2 snRNAs in A. castellanii, P. ramorum, P. sojae and their human homologues. Also shown is experimental evidence for expression of A. castellanii U12 snRNA and annotation of genomic sequences encoding all the snRNAs identified in this study
Oligonucleotide primers and strategy used to search for minor spliceosomal introns and Phytophthora U11 snRNA homologues. Contains the sequences and descriptions of oligonucleotide primers used in this study. Specific search parameters used for the identification of U12-type introns and Phytophthora U11 snRNAs are described
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Biology Direct (2012)