An uncapped RNA suggests a model for <i>Caenorhabditis elegans</i> polycistronic pre-mRNA processing

Liu, Yingmiao; Kuersten, Scott; Huang, Tao; Larsen, Alison; MacMorris, Margaret; Blumenthal, Thomas

doi:10.1261/rna.2128903

Cited by 19 publications

(20 citation statements)

References 28 publications

(26 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Overall, these results provide evidence that RNAPII is pausing at all 3 0 ends in operons, suggesting that internal 3 0 ends in operons is primed for termination but is prevented by a mechanism other than affecting RNAPII or CstF recruitment. Liu et al, 65 proposed that RNA-PII is prevented from terminating at internal 3 0 ends by a factor bound to the Ur element, a sequence between operon genes required for SL2 trans-splicing. This factor could temporarily block the passage of the XRN-2 exonuclease until SL2 trans-splicing provides a cap to the downstream RNA.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Localization of RNAPII and 3′ end formation factor CstF subunits onC. elegansgenes and operons

2016

Self Cite

View full text Add to dashboard Cite

Transcription termination is mechanistically coupled to pre-mRNA 3 0 end formation to prevent transcription much beyond the gene 3 0 end. C. elegans, however, engages in polycistronic transcription of operons in which 3 0 end formation between genes is not accompanied by termination. We have performed RNA polymerase II (RNAPII) and CstF ChIP-seq experiments to investigate at a genome-wide level how RNAPII can transcribe through multiple poly-A signals without causing termination. Our data shows that transcription proceeds in some ways as if operons were composed of multiple adjacent single genes. Total RNAPII shows a small peak at the promoter of the gene cluster and a much larger peak at 3 0 ends. These 3 0 peaks coincide with maximal phosphorylation of Ser2 within the C-terminal domain (CTD) of RNAPII and maximal localization of the 3 0 end formation factor CstF. This pattern occurs at all 3 0 ends including those at internal sites in operons where termination does not occur. Thus the normal mechanism of 3 0 end formation does not always result in transcription termination. Furthermore, reduction of CstF50 by RNAi did not substantially alter the pattern of CstF64, total RNAPII, or Ser2 phosphorylation at either internal or terminal 3 0 ends. However, CstF50 RNAi did result in a subtle reduction of CstF64 binding upstream of the site of 3 0 cleavage, suggesting that the CstF50/CTD interaction may facilitate bringing the 3 0 end machinery to the transcription complex.

show abstract

Section: Discussionmentioning

confidence: 99%

“…A likely possibility is that the SL2 trans-splicing that co-occurs with 3 0 end formation provides a 5 0 cap to the downstream RNA, so the XRN2 exonuclease cannot act to cause termination by the usual torpedo mechanism. 65 CstF50 plays a role in CstF64 localization upstream of the cleavage site…”

Section: Cstf Distribution On C Elegans Operonsmentioning

confidence: 99%

Localization of RNAPII and 3′ end formation factor CstF subunits onC. elegansgenes and operons

2016

Self Cite

View full text Add to dashboard Cite

show abstract

“…Sequences required for 39-end cleavage are also required for SL2 transsplicing. Following cleavage catalyzed by the cleavage and polyadenylation specificity factor (CPSF) bound at the AAUAAA just 59 of the cleavage site and cleavage stimulatory factor (CstF) bound to U-rich sequences in the vicinity of the cleavage site (Salisbury et al 2006), the 39 portion of the pre-mRNA is degraded exonucleolytically 59 to 39 until the nuclease reaches a block, possibly a bound protein (Liu et al 2003). CstF has been shown to interact with the SL2 snRNP, and mutations within the third stem-loop of SL2 RNA that eliminate this binding prevent its trans-splicing at operon trans-splice sites (Evans et al 2001).…”

Section: Introductionmentioning

confidence: 99%

A novel family of C. elegans snRNPs contains proteins associated with trans-splicing

MacMorris

Kumar

Lasda

et al. 2007

RNA

Self Cite

View full text Add to dashboard Cite

In many Caenorhabditis elegans pre-mRNAs, the RNA sequence between the 59 cap and the first 39 splice site is replaced by trans-splicing a short spliced leader (SL) from the Sm snRNP, SL1. C. elegans also utilizes a similar Sm snRNP, SL2, to trans-splice at sites between genes in polycistronic pre-mRNAs from operons. How do SL1 and SL2 snRNPs function in different contexts? Here we show that the SL1 snRNP contains a complex of SL75p and SL21p, which are homologs of novel proteins previously reported in the Ascaris SL snRNP. Interestingly, we show that the SL2 snRNP does not contain these proteins. However, SL75p and SL26p, a paralog of SL21p, are components of another Sm snRNP that contains a novel snRNA species, Sm Y. Knockdown of SL75p is lethal. However, knockdown of either SL21p or SL26p alone leads to cold-sensitive sterility, whereas knockdown of both SL21p and SL26p is lethal. This suggests that these two proteins have overlapping functions even though they are associated with different classes of snRNP. These phenotypic relationships, along with the association of SL26p with SL75p, imply that, like the SL1 RNA/Sm/SL75p/SL21p complex, the Sm Y/Sm/SL75p/SL26p complex is associated with trans-splicing.

show abstract

“…Additionally, the stem or anything bound to it may block 59-to-39 exonucleolytic RNA degradation following cleavage at the 39 end of the upstream gene. Supporting this, the laboratory (Liu et al 2003) has reported previously the existence of a ''Ur RNA'' from the gpd-2/gpd-3 transgenic operon with a 59 end 4 nt upstream of the Ur element stem, a location dependent on the position of the Ur element.…”

Section: The Stem-loop Of the Ur Elementmentioning

confidence: 65%

“…Downstream operon genes are not cotranscriptionally capped; therefore, upstream gene 39 end cleavage would leave an unprotected 59 end on the downstream premRNA. The uncapped mRNA would likely be rapidly degraded 59 to 39 (Liu et al 2003). Linking the cleavage that generates the uncapped 59 RNA end with trans-splicing that provides a 59 TMG cap would ensure protection of the downstream mRNA, as well as prevent transcription termination that is associated with 39 end formation.…”

Section: Concurrent Transcription Is Not Required For Sl2 Specificitymentioning

confidence: 99%

Polycistronic pre-mRNA processing in vitro: snRNP and pre-mRNA role reversal in trans-splicing

Lasda

Allen²,

Blumenthal³

2010

Genes Dev.

Self Cite

View full text Add to dashboard Cite

Spliced leader (SL) trans-splicing in Caenorhabditis elegans attaches a 22-nucleotide (nt) exon onto the 59 end of many mRNAs. A particular class of SL, SL2, splices mRNAs of downstream operon genes. Here we use an embryonic extract-based in vitro splicing system to show that SL2 specificity information is encoded within the polycistronic pre-mRNA, and that trans-splicing specificity is recapitulated in vitro. We define an RNA sequence required for SL2 trans-splicing, the U-rich (Ur) element, through mutational analysis and bioinformatics as a short stem-loop followed by a sequence motif, UAYYUU, located~50 nt upstream of the trans-splice site. Furthermore, this element is predicted in intercistronic regions of numerous operons of C. elegans and other species that use SL2 trans-splicing. We propose that the UAYYUU motif hybridizes with the 59 splice site on the SL2 RNA to recruit the SL to the pre-mRNA. In this way, the UAYYUU motif in the pre-mRNA would serve an analogous function to the similar sequence in the U1 snRNA, which binds to the 59 splice site of introns, effectively reversing the roles of snRNP and pre-mRNA in trans-splicing. In spliced leader (SL) trans-splicing, a short leader sequence is transferred from the 59 end of an SL RNA onto the first exon of a pre-mRNA (for review, see Nilsen 1993;Blumenthal 2005;Hastings 2005). This joins two discontinuous RNA sequences, and provides a common 59 sequence to many mRNAs. SL trans-splicing was identified first in trypanosomes and subsequently in numerous other organisms, ranging from protists to primitive chordates, and including roundworms, flatworms, arthropods, and cnidarians. However, it is not found in plants, A different form of trans-splicing has been reported in flies (Mongelard et al. 2002), and also recently in worms (Fischer et al. 2008). In these instances, exons of separate pre-mRNAs, neither one an SL RNA, are spliced together to form a mature mRNA containing portions of both premRNAs. Additionally, there have been numerous reports of a similar process in mammalian cells (Akiva et al. 2006) and plants (Zhang et al. 2010), where normally separate mRNAs are ''accidentally'' trans-spliced together. This sort of splicing has been implicated recently in cancers associated with translocations (Li et al. 2008).SL trans-splicing and cis-splicing (intron removal) are similar, and trans-splicing likely evolved from cis-splicing (Blumenthal 2004(Blumenthal , 2005. The splice site on the SL RNA closely matches the 59 splice site (59ss) consensus sequence, and the trans-splice site of the pre-mRNA has the same consensus sequence as intronic 39ss (Kent and Zahler 2000;Blumenthal 2005). In Caenorhabditis elegans, trans-splicing is signaled by a 39ss without an upstream 59ss. Artificial trans-splice sites can be created from cis-splice sites by removing an upstream 59ss (Conrad et al. 1991(Conrad et al. , 1995Maroney et al. 2000;Boukis and Bruzik 2001), and a trans-splice site can be made to cis-splice instead by insertion of a 59ss into the outron, the intr...

show abstract

An uncapped RNA suggests a model for Caenorhabditis elegans polycistronic pre-mRNA processing

Cited by 19 publications

References 28 publications

Localization of RNAPII and 3′ end formation factor CstF subunits onC. elegansgenes and operons

Localization of RNAPII and 3′ end formation factor CstF subunits onC. elegansgenes and operons

A novel family of C. elegans snRNPs contains proteins associated with trans-splicing

Polycistronic pre-mRNA processing in vitro: snRNP and pre-mRNA role reversal in trans-splicing

Contact Info

Product

Resources

About