A dominant role for meiosis-specific 3' RNA processing in controlling expression of a fission yeast cyclin gene.

Meiotic gene regulation provides a rich source of insight into mechanisms of temporal control during development. We previously reported that accumulation of many meiotic mRNAs in fission yeast is governed by changes in 3' RNA processing and elucidated the molecular basis of this regulatory mechanism for an early meiotic gene. Here, we report that cleavage/polyadenylation is also the nexus of negative control for middle meiotic genes. Parallel profiles of splicing and polyadenylation are observed over a meiotic time course for both rem1 and spo4 but not for a constitutive control gene. Nevertheless, polyadenylation of rem1 transcripts is restricted to meiosis by a splicing-independent mechanism. Through systematic sequence substitutions, we identified a negative control region (NCR) located upstream of the rem1 transcription start site and found that it is required to block 3' RNA processing in proliferating cells. Ablation of the NCR relieves inhibition regardless of whether the intron is present, absent, or carries splice site mutations. Consistent with the previous report of a polypeptide encoded by the first exon of rem1, we discovered a second 3' processing site just downstream from the 5' splice site. Polyadenylation within the intron is activated concurrent with the downstream site during meiosis, is controlled by the NCR, and is enhanced when splicing is blocked via 5' junction or branch point mutations. Taken together, these data suggest a novel regulatory mechanism in which a 5' element modulates the dynamic interplay between splicing and polyadenylation.

A meiotic gene regulatory cascade driven by alternative fates for newly synthesized transcripts.

To determine the relative importance of transcriptional regulation versus RNA processing and turnover during the transition from proliferation to meiotic differentiation in the fission yeast Schizosaccharomyces pombe, we analyzed temporal profiles and effects of RNA surveillance factor mutants on expression of 32 meiotic genes. A comparison of nascent transcription with steady-state RNA accumulation reveals that the vast majority of these genes show a lag between maximal RNA synthesis and peak RNA accumulation. During meiosis, total RNA levels parallel 3' processing, which occurs in multiple, temporally distinct waves that peak from 3 to 6 h after meiotic induction. Most early genes and one middle gene, mei4, share a regulatory mechanism in which a specialized RNA surveillance factor targets newly synthesized transcripts for destruction. Mei4p, a member of the forkhead transcription factor family, in turn regulates a host of downstream genes. Remarkably, a spike in transcription is observed for less than one-third of the genes surveyed, and even these show evidence of RNA-level regulation. In aggregate, our findings lead us to propose that a regulatory cascade driven by changes in processing and stability of newly synthesized transcripts operates alongside the well-known transcriptional cascade as fission yeast cells enter meiosis.

A complex gene regulatory mechanism that operates at the nexus of multiple RNA processing decisions.

Expression of crs1 pre-mRNA, encoding a meiotic cyclin, is blocked in actively growing fission yeast cells by a multifaceted mechanism. The most striking feature is that in vegetative cells, crs1 transcripts are continuously synthesized but are targeted for degradation rather than splicing and polyadenylation. Turnover of crs1 RNA requires the exosome, as do previously described nuclear surveillance and silencing mechanisms, but does not involve a noncanonical poly(A) polymerase. Instead, crs1 transcripts are targeted for destruction by a factor previously implicated in turnover of meiotic RNAs in growing cells. Like exosome mutants, mmi1 mutants splice and polyadenylate vegetative crs1 transcripts. Two regulatory elements are located at the 3' end of the crs1 gene, consistent with the increased accumulation of spliced RNA in polyadenylation factor mutants. This highly integrated regulatory strategy may ensure a rapid response to adverse conditions, thereby guaranteeing survival.