Abstract
BackgroundTwo spliceosomal intron types co-exist in eukaryotic precursor mRNAs and are excised by distinct U2-dependent and U12-dependent spliceosomes. In the diplomonad Giardia lamblia, small nuclear (sn) RNAs show hybrid characteristics of U2- and U12-dependent spliceosomal snRNAs and 5 of 11 identified remaining spliceosomal introns are trans-spliced. It is unknown whether unusual intron and spliceosome features are conserved in other diplomonads.ResultsWe have identified spliceosomal introns, snRNAs and proteins from two additional diplomonads for which genome information is currently available, Spironucleus vortens and Spironucleus salmonicida, as well as relatives, including 6 verified cis-spliceosomal introns in S. vortens. Intron splicing signals are mostly conserved between the Spironucleus species and G. lamblia. Similar to ‘long’ G. lamblia introns, RNA secondary structural potential is evident for ‘long’ (> 50 nt) Spironucleus introns as well as introns identified in the parabasalid Trichomonas vaginalis. Base pairing within these introns is predicted to constrain spatial distances between splice junctions to similar distances seen in the shorter and uniformly-sized introns in these organisms. We find that several remaining Spironucleus spliceosomal introns are ancient. We identified a candidate U2 snRNA from S. vortens, and U2 and U5 snRNAs in S. salmonicida; cumulatively, illustrating significant snRNA differences within some diplomonads. Finally, we studied spliceosomal protein complements and find protein sets in Giardia, Spironucleus and Trepomonas sp. PC1 highly- reduced but well conserved across the clade, with between 44 and 62 out of 174 studied spliceosomal proteins detectable. Comparison with more distant relatives revealed a highly nested pattern, with the more intron-rich fornicate Kipferlia bialata retaining 87 total proteins including nearly all those observed in the diplomonad representatives, and the oxymonad Monocercomonoides retaining 115 total proteins including nearly all those observed in K. bialata.ConclusionsComparisons in diplomonad representatives and species of other closely-related metamonad groups indicates similar patterns of intron structural conservation and spliceosomal protein composition but significant divergence of snRNA structure in genomically-reduced species. Relative to other eukaryotes, loss of evolutionarily-conserved snRNA domains and common sets of spliceosomal proteins point to a more streamlined splicing mechanism, where intron sequences and structures may be functionally compensating for the minimalization of spliceosome components.
Highlights
Two spliceosomal intron types co-exist in eukaryotic precursor mRNAs and are excised by distinct U2dependent and U12-dependent spliceosomes
Spliceosomal introns in ribosomal protein (RP) and non-RP genes in S. vortens Only 11 spliceosomal introns have been identified in the diplomonad Giardia lamblia [1, 16, 19,20,21,22,23], revealing both a remarkable paucity of spliceosomal introns and proportionally large number of trans-spliced introns in this organism
Our search strategy employed the conserved G. lamblia 5′ splice site (SS) sequence ‘VTATGTT’ and fused branch point (BP) and 3′ SS sequence ‘VCTRACACRCAG’ (‘R’ is a purine; ‘V’ is an A, C, or G nucleotide) [20], but these searches did not identify any introns in S. vortens
Summary
Two spliceosomal intron types co-exist in eukaryotic precursor mRNAs and are excised by distinct U2dependent and U12-dependent spliceosomes. In the diplomonad Giardia lamblia, small nuclear (sn) RNAs show hybrid characteristics of U2- and U12-dependent spliceosomal snRNAs and 5 of 11 identified remaining spliceosomal introns are trans-spliced. It is unknown whether unusual intron and spliceosome features are conserved in other diplomonads. Eukaryotic nuclear genomes contain spliceosomal introns which divide protein-coding sequences into separate exons. Some intron-poor species contain only a few introns per genome [1, 2] while some intronrich species have on average several introns per kilobase of gene sequence [3]. A few previous studies have revealed phylogenetic diversity of the spliceosomal machinery itself, less is known about the evolution of the spliceosome than about the evolution of introns
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