5' splice site selection in yeast: genetic alterations in base-pairing with U1 reveal additional requirements.
Using a strategy of compensatory nucleotide changes between yeast U1 and a 5' splice site, we have analyzed the contribution of base-pairing to the efficiency and fidelity of pre-mRNA splicing in vivo. Watson-Crick base-pairing interactions with U1 can be demonstrated at intron positions 1 and 5 but not at position 4. Moreover, restoration of the ability to pair with U1 is not sufficient to restore activity in the second step of splicing to intron position 1 mutants. Finally, in contrast to recent observations in mammalian systems, we find that the precise position of 5' splice site cleavage is not determined solely by the base-pairing interaction with U1. Rather, the presence of a G residue at position 5 is required for the correct localization of the nucleolytic event. Taken together, these results indicate that the demands for 5' splice site selection and utilization are more complex than a simple maximization of Watson-Crick interactions with U1.
Siliciano
P G
PG
Department of Biochemistry and Biophysics, University of California, San Francisco 94143.
Guthrie
C
C
D001482:Base Composition; D003001:Cloning, Molecular; D007438:Introns; D009154:Mutation; D012326:RNA Splicing; D012342:RNA, Small Nuclear / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D014170:Transformation, Genetic
D012342:RNA, Small Nuclear