A bidirectional terminator, predicted to be shared with the converent cueO mRNA, is at the end of a 187 nt 3' UTR for gcd.
The predicted 187 nt long 3' UTR for gcd contains the RIP10 REP element. This EcoGene annotation of a 187 nt 3' UTR for gcd is only partially supported by the highthroughput data of Conway et al., who found three 3' ends within this annotated 3' UTR. (Conway, 2014).
Conway et al. observe a convergent transcript's 3' end at this bidirectional terminator (Conway, 2014). However, the Conway-observed 3' end is in the tetraloop, which is unusual. Conway et al. state that the 30 nts just prior to their observed 3' end contain a predicted factor-independent terminator as confirmed by TransTermHP, but this claim could not be validated using mfold, which predicts no secondary structures predicting a terminator, in contrast to the excellent mfold prediction of a bidirectional terminator. Perhaps the loop was cut by an RNase during mRNA extraction prior to the RNA-Seq step. Until this issue is resolved by validation of the Conway-observed highthroughput 3' end, the gcd mRNA annotated in EcoGene will extend to the end of the EcoGene-predicted bidirectional terminator.
Conway et al. observe two 3' ends prior to reaching this terminator. Immediately before the RIP10 element is a weakly predicted terminator, that could alternatively be a pause site or an RNase III cleavage site, that would create a 39 nt 3' UTR. Another 3' end was observed in the IHF sequence of RIP10, but at a different location than the convergent cueO mRNA 3' end observed in RIP10's IHF sequence (see cueO 3' UTR TopicPage). No secondary structure is predicted for the RNA preceding this 3' end.
Convergent transcripts are abundant and present special problems for transcription termiantion as some of the convergent genes actually overlap and have no intergenic region and thus must terminate in the antisense RNA of the convergent gene's codng region. Indeed Conway et al. found that 276 convergent mRNAs terminate well within the converging gene, creating an average overlap (dsRNA) of 286 nts; however they also found 94 converging gene pairs, like cueO-gcd, that do not read through into each other (Conway, 2014). The cueO-gcd intergenic region has two 3' ends for cueO and three for gcd. Although they may not all be termination events (perhaps some long pauses or an RNase III cleavage), the presence of RIP10, a predicted bidirectinal terminator, and a possible RNase III cleavage site may indicate that antisense mRNA for these genes is deleterious to cell growth or survival and has been selected against. Another consideration is the fact that convergent transcription can introduced positive supercoils that may need to be actively rewound to negative supercoils by DNA gyrase. REP elements have been found to bind DNA gyrase so possibly RIP10 serves this purpose. The IHF site might stabilize the REP-gyrase complex, which might also be responsible for the generation of 3' ends from pause sites in this region when disrupted. cueO is predicted to be a foreign gene and gcd is predicted to be a native gene; perhaps when cueO was transmitted to E. coli the sudden juxaposition of these converging transcripts was problematic, leading to the subsequent insertion of RIP10 or the other intergenes to block the deleterious antisense mRNAs from inpinging upon mRNA function or stability.
Bibliography (1 total) : Review Only   Up
Conway T, Creecy JP, Maddox SM, Grissom JE, Conkle TL, Shadid TM, Teramoto J, San Miguel P, Shimada T, Ishihama A, Mori H, Wanner BL (2014) Unprecedented high-resolution view of bacterial operon architecture revealed by RNA sequencing. MBio 5:e01442-14