THE PLANT CELL, Vol 8, Issue 12 2295-2307, Copyright © 1996 by American Society of Plant Biologists
Interactions across Exons Can Influence Splice Site Recognition in Plant Nuclei
A. J. McCullough, C. E. Baynton and M. A. Schuler
Department of Plant Biology, University of Illinois, Urbana, Illinois 61801
In vivo analyses of cis-acting sequence requirements for pre-mRNA splicing
in tobacco nuclei have previously demonstrated that the 5[prime] splice
sites are selected by their position relative to AU-rich elements within
plant introns and by their degree of complementarity to the U1 small
nuclear RNA. To determine whether the presence of adjacent introns affects
5[prime] splice site recognition in plant nuclei, we have analyzed the in
vivo splicing patterns of two-intron constructs containing 5[prime] splice
site mutations in the second intron. These experiments indicated that the
splice site selection patterns in plant nuclei are defined primarily by
sequences within the intron (intron definition) and secondarily by weak
interactions across exons (exon definition). The effects of these secondary
interactions became evident only when mutations in the downstream 5[prime]
splice site decreased its functionality and differed depending on the
availability of cryptic splice sites close to the mutant site. In
[beta]-conglycinin chimeric transcripts containing multiple cryptic
5[prime] splice sites, the presence of an intact upstream intron
significantly increased splicing at the downstream 5[prime] splice sites in
a polar fashion without activating exon skipping. In a natural
[beta]-conglycinin transcript, which does not contain cryptic 5[prime]
splice sites, mutation of the first nucleotide of the downstream intron
activated an array of noncanonical 5[prime] and 3[prime] splice sites and
some exon skipping.
