A conditional U5 snRNA mutation affecting pre-mRNA splicing and nuclear pre-mRNA retention identifies SSD1/SRK1 as a general splicing mutant suppressor.
A combination of point mutations disrupting both stem 1 and stem 2 of U5 snRNA (U5AI) was found to confer a thermosensitive phenotype in vivo. In a strain expressing U5AI, pre-mRNA splicing was blocked before the first step through an inability of the mutant U5 snRNA to efficiently associate with the U4/U6 di-snRNP. Formation of early splicing complexes was not affected in extracts prepared from U5 snRNA mutant cells, while the capacity of these extracts to splice a pre-mRNA in vitro was greatly diminished. In addition, significant levels of a translation product derived from intron containing pre-mRNAs could be detected in vivo. The SSD1/SRK1 gene was identified as a multi-copy suppressor of the U5AI snRNA mutant. Single copy expression of SSD1/SRK1 was sufficient to suppress the thermosensitive phenotype, and high copy expression partially suppressed the splicing and U4/U6.U5 tri-snRNP assembly pheno-types. SSD1/SRK1 also suppressed thermosensitive mutations in the Prp18p and U1-70K proteins, while inhibiting growth of the cold sensitive U1-4U snRNA mutant at 30 degrees C. Thus we have identified SSD1/SRK1 as a general suppressor of splicing mutants.
Luukkonen
B G
BG
European Molecular Biology Laboratory, Meyerhofstrasse 1, D-69117 Heidelberg, Germany.
Séraphin
B
B
D001483:Base Sequence; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics; D016153:Genes, Suppressor / Q000235:genetics* / Q000502:physiology*; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D010641:Phenotype; D012322:RNA Precursors / Q000737:chemistry / Q000235:genetics*; D012326:RNA Splicing; D012342:RNA, Small Nuclear / Q000737:chemistry / Q000235:genetics*; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000378:metabolism; D013489:Suppression, Genetic; D013696:Temperature
D005656:Fungal Proteins; D012322:RNA Precursors; D012342:RNA, Small Nuclear; D017414:Ribonucleoprotein, U4-U6 Small Nuclear