Poly(A) site choice during mRNA 3'-end formation in the Schizosaccharomyces pombe wos2 gene

In the fission yeast Schizosaccharomyces pombe, the wos2 gene encodes p23, a highly conserved protein which functions as a co-chaperone for the heat shock protein Hsp90. This p23 protein binds to Hsp90, but its activities and regulatory mechanisms are still unclear. Northern analysis has shown that the wos2 gene produces three transcripts of about 1.1, 0.9 and 0.8 kb, which are expressed differentially depending on the growth temperature. The largest and the smallest transcripts were most abundant at 25 degrees C, whereas the 0.9-kb transcript predominated at 37 degrees C. A time-course analysis indicated that this 0.9-kb species rapidly increased in abundance after a shift from 25 degrees C to 37 degrees C, reaching a maximum after 15 min. A shift back to 25 degrees C resulted in a decline in the amount of this transcript, albeit at a slower rate. Expression analysis of wos2:ura4 and nmt1:wos2 constructs showed that the 3' untranslated region of wos2 alone directs the formation of these multiple, discrete wos2 mRNAs. Sequence analysis of cDNAs derived from these mRNAs showed that the use of different polyadenylation sites results in the production of the three differently sized wos2 transcripts. In the case of the 0.9- and 0.8-kb mRNA species, these sites lie in a predicted hairpin loop in the mRNA, suggesting that polyadenylation signals in wos2 transcripts may be mediated by RNA secondary structure. The possibility that differential thermal stability of these hairpin structures could influence polyadenylation site choice during formation of the 3'-ends of the mRNAs is discussed.