Abstract
The 2,2,7-trimethylguanosine (TMG) cap is one of the first identified modifications on eukaryotic RNAs. TMG, synthesized by the conserved Tgs1 enzyme, is abundantly present on snRNAs essential for pre-mRNA splicing. Results from ex vivo experiments in vertebrate cells suggested that TMG ensures nuclear localization of snRNAs. Functional studies of TMG using tgs1 mutations in unicellular organisms yield results inconsistent with TMG being indispensable for either nuclear import or splicing. Utilizing a hypomorphic tgs1 mutation in Drosophila, we show that TMG reduction impairs germline development by disrupting the processing, particularly of introns with smaller sizes and weaker splice sites. Unexpectedly, loss of TMG does not disrupt snRNAs localization to the nucleus, disputing an essential role of TMG in snRNA transport. Tgs1 loss also leads to defective 3’ processing of snRNAs. Remarkably, stronger tgs1 mutations cause lethality without severely disrupting splicing, likely due to the preponderance of TMG-capped snRNPs. Tgs1, a predominantly nucleolar protein in Drosophila, likely carries out splicing-independent functions indispensable for animal development. Taken together, our results suggest that nuclear import is not a conserved function of TMG. As a distinctive structure on RNA, particularly non-coding RNA, we suggest that TMG prevents spurious interactions detrimental to the function of RNAs that it modifies.
Highlights
Post-transcriptional modifications have been increasingly recognized as essential for the function of RNA molecules
Chemical modifications on RNAs regulate their function in vivo
TMG is abundantly present on most members of small nuclear RNA that are essential for pre-mRNA
Summary
Post-transcriptional modifications have been increasingly recognized as essential for the function of RNA molecules. TMG, which is different from the m7G cap on mRNA molecules, is present on many species of non-coding RNAs such as small nuclear RNA U1, U2, U4 and U5, essential for pre-mRNA splicing (spliceosomal snRNAs) (reviewed in [2,3,4,5]), and U7 snRNA, essential for histone RNA processing [6]. It is present on some of the small nucleolar RNAs (snoRNAs) that function in the processing of ribosomal RNAs [7]. TMG has been shown to promote the expression of viral mRNAs from HIV [12], and the biogenesis of quiescence-induced miRNAs in human cells [13]
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