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Poly(A) signals located near the 5' end of genes are silenced by a general mechanism that prevents premature 3'-end processing.

ABSTRACT: Poly(A) signals located at the 3' end of eukaryotic genes drive cleavage and polyadenylation at the same end of pre-mRNA. Although these sequences are expected only at the 3' end of genes, we found that strong poly(A) signals are also predicted within the 5' untranslated regions (UTRs) of many Drosophila melanogaster mRNAs. Most of these 5' poly(A) signals have little influence on the processing of the endogenous transcripts, but they are very active when placed at the 3' end of reporter genes. In investigating these unexpected observations, we discovered that both these novel poly(A) signals and standard poly(A) signals become functionally silent when they are positioned close to transcription start sites in either Drosophila or human cells. This indicates that the stage when the poly(A) signal emerges from the polymerase II (Pol II) transcription complex determines whether a putative poly(A) signal is recognized as functional. The data suggest that this mechanism, which probably prevents cryptic poly(A) signals from causing premature transcription termination, depends on low Ser2 phosphorylation of the C-terminal domain of Pol II and inefficient recruitment of processing factors.

SUBMITTER: Guo J

PROVIDER: S-EPMC3028650 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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Poly(A) signals located near the 5' end of genes are silenced by a general mechanism that prevents premature 3'-end processing.

Guo Jiannan J Garrett Matthew M Micklem Gos G Brogna Saverio S

Molecular and cellular biology 20101206 4

Poly(A) signals located at the 3' end of eukaryotic genes drive cleavage and polyadenylation at the same end of pre-mRNA. Although these sequences are expected only at the 3' end of genes, we found that strong poly(A) signals are also predicted within the 5' untranslated regions (UTRs) of many Drosophila melanogaster mRNAs. Most of these 5' poly(A) signals have little influence on the processing of the endogenous transcripts, but they are very active when placed at the 3' end of reporter genes. ...[more]

PMID: 21135120

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Project description:Evolutionarily conserved NusG protein enhances bacterial RNA polymerase processivity but can also promote transcription termination by binding to, and stimulating the activity of, Rho factor. Rho terminates transcription upon anchoring to cytidine-rich motifs, the so-called Rho utilization sites (Rut) in nascent RNA. Both NusG and Rho have been implicated in the silencing of horizontally-acquired A/T-rich DNA by nucleoid structuring protein H-NS. However, the relative roles of the two proteins in H-NS-mediated gene silencing remain incompletely defined. In the present study, a Salmonella strain carrying the nusG gene under the control of an arabinose-inducible repressor was used to assess the genome-wide response to NusG depletion. Results from two complementary approaches, i) screening lacZ protein fusions generated by random transposition and ii) transcriptomic analysis, converged to show that loss of NusG causes massive upregulation of Salmonella pathogenicity islands (SPIs) and other H-NS-silenced loci. A similar, although not identical, SPI-upregulated profile was observed in a strain with a mutation in the rho gene, Rho K130Q. Surprisingly, Rho mutation Y80C, which affects Rho's primary RNA binding domain, had either no effect or made H-NS-mediated silencing of SPIs even tighter. Thus, while corroborating the notion that bound H-NS can trigger Rho-dependent transcription termination in vivo, these data suggest that H-NS-elicited termination occurs entirely through a NusG-dependent pathway and is less dependent on Rut site binding by Rho. We provide evidence that through Rho recruitment, and possibly through other still unidentified mechanisms, NusG prevents pervasive transcripts from elongating into H-NS-silenced regions. Failure to perform this function causes the feedforward activation of the entire Salmonella virulence program. These findings provide further insight into NusG/Rho contribution in H-NS-mediated gene silencing and underscore the importance of this contribution for the proper functioning of a global regulatory response in growing bacteria. The complete set of transcriptomic data is freely available for viewing through a user-friendly genome browser interface.

Dataset Information

Poly(A) signals located near the 5' end of genes are silenced by a general mechanism that prevents premature 3'-end processing.

Publications

Poly(A) signals located near the 5' end of genes are silenced by a general mechanism that prevents premature 3'-end processing.

Similar Datasets

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