Alternative polyadenylation in the regulation and dysregulation of gene expression.

Transcriptional control shapes a cell's transcriptome composition, but it is RNA processing that refines its expression. The untranslated regions (UTRs) of mRNA are hotspots for regulatory control. Features in these can impact mRNA stability, localisation and translation. Here we describe how alternative cleavage and polyadenylation can change mRNA fate by changing the length of its 3'UTR.

Understanding the regulation of coding and noncoding transcription in cell populations.

Whole transcriptome analyses have unveiled the uncomfortable truth that we know less about how transcription is regulated then we thought. In addition to its role in classic promoter-driven transcription of coding RNA, it is now clear that RNA Pol II also drives abundant expression of noncoding RNA. For the majority of this the functional significance remains unclear. Moreover, its regulation and impact are hard to predict because it often proceeds in unexpected ways from cryptic promoters, including by driving convergent antisense transcription from within 3' UTRs. This review suggests that its time to rethink how we envisage gene expression by inclusion of the regulatory architecture of the full genetic locus, and expanding our thinking to encompass the fact that we generally study cells within heterogeneous populations.

Using Klenow-mediated extension to measure poly(A)-tail length and position in the transcriptome.

The poly(A)-tail that terminates most mRNA and many noncoding RNA is a convenient "hook" to isolate mRNA. However the length of this tail and its position within the primary RNA transcript can also hold diagnostic value for RNA metabolism. In general, mRNA with a long poly(A)-tail is well translated, whereas a short poly(A)-tail can indicate translational silencing. A short poly(A)-tail is also appended to RNA-decay intermediates via the TRAMP complex. A number of approaches have been developed to measure the length and position of the poly(A)-tail. Here, we describe a simple method to "tag" adenylated RNA using the native function of DNA polymerase I to extend an RNA primer on a DNA template in second-strand DNA synthesis. This function can be harnessed as a means to purify, visualize, and quantitate poly(A)-dynamics of individual RNA and the transcriptome en masse.