Heat shock affects 5' splice site selection, cleavage and ligation of CAD pre-mRNA in hamster cells, but not its packaging in InRNP particles.
The effect of heat shock on the packaging and splicing of nuclear CAD pre-mRNA, a transcript expressed constitutively from a non heat-inducible promoter, was studied in vivo in Syrian hamster cells. While mild heat shock did not affect significantly the packaging of CAD RNA in 200S InRNP particles, it caused perturbation to splicing. First, the heat shock inhibited splicing of CAD pre-mRNA. Second, it affected 5' splice site selection by activating cleavage at a cryptic 5' splice site; yet ligation of the cryptic exon to the downstream proximal exon was not observed. Base complementarities of the cryptic site with U1, U5, or U6 snRNAs are comparable, or even better, than those with the neighboring normal site. Hence, the exclusion of the cryptic site under normal growth conditions cannot be attributed to weaker base pairing with these snRNAs. On the other hand, these results imply the involvement of a heat labile factor in the selection of the 5' cleavage site. The exclusion of the cryptic site at 37 degrees C and the aborted splicing at this site after heat shock may also be explained by a proposed nuclear checking mechanism that detects in-frame stop codons upstream of the 5' splice site, and aborts splicing at such sites to prevent the production of a defective message.
Miriami
E
E
Department of Genetics, Hebrew University of Jerusalem, Givat Ram, Israel.
Sperling
J
J
Sperling
R
R
D000818:Animals; D001221:Aspartate Carbamoyltransferase / Q000235:genetics; D001483:Base Sequence; D002223:Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing) / Q000235:genetics; D002467:Cell Nucleus / Q000378:metabolism; D016384:Consensus Sequence; D006224:Cricetinae; D004080:Dihydroorotase / Q000235:genetics; D006358:Hot Temperature; D008647:Mesocricetus; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012322:RNA Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012326:RNA Splicing; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000378:metabolism*
D009097:Multienzyme Complexes; D012322:RNA Precursors; D012333:RNA, Messenger; D012261:Ribonucleoproteins; C019820:carbamoyl phosphate synthase-aspartatecarbamoyl transferase complex; D001221:Aspartate Carbamoyltransferase; D004080:Dihydroorotase; D002223:Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing)