AN INTRONIC (A U)GGG REPEAT ENHANCES THE SPLICING OF AN ALTERNATIVE INTRON OF THE CHICKEN BETA-TROPOMYOSIN PRE-MESSENGER-RNA/

Computer analysis of human intron sequences have revealed a 50 nucleot ide (nt) GC-rich region down-stream of the 5' splice site; the trinucl eotide GGG occurs almost four times as frequently as it would in a ran dom sequence, The 5' part of a beta-tropomyosin intron exhibits six re petitions of the motif (A/U)GGG, In order to test whether these motifs play a role in the splicing process we have mutated some or all of th em, Mutated RNAs show a lower in vitro splicing efficiency when compar ed with the wild-type, especially when all six motifs are mutated (>70 % inhibition), Assembly of the spliceosome complex B and, to a lesser extent, of the pre-spliceosome complex A also appears to be strongly a ffected by this mutation, A 55 kDa protein within HeLa cell nuclear ex tract is efficiently cross-linked to the G-rich region, This protein i s present in the splicing complexes and its cross-linking to the pre-m RNA requires the presence of one or several snRNP, Altogether our resu lts suggest that the G-rich sequences present in the 5' part of intron s may act as an enhancer of the splicing reaction at the level of spli ceosome assembly.