Determinants for cap trimethylation of the U2 small nuclear RNA are not conserved between Trypanosoma brucei and higher eukaryotic organisms

In most eukaryotic organisms the U2 small nuclear RNA (snRNA) gene is trans cribed by RNA polymerase II to generate a primary transcript with a 5' term inal 7-methylguanosine cap structure, Following nuclear export, the U2 snRN A is assembled into a core ribonucleoprotein particle (RNP), This involves binding a set of proteins that are shared by spliceosomal snRNPs to the hig hly conserved Sm site, Prior to nuclear import, the snRNA-(guanosine-N2)-me thyltransferase appears to interact with the core RNP and hypermethylates t he cap structure to 2,2,7-trimethylguanosine (m(3)G), In the protist parasi te Trypanosoma brucei, U-snRNAs are complexed with a set of common proteins that are analogous to eukaryotic Sm antigens but do not have a highly cons erved Sm sequence motif, and most U-snRNAs are synthesised by RNA polymeras e iii. Here, we examined the determinants for m(3)G cap formation in T. bru cei by expressing mutant U2 snRNAs in vivo and assaying trimethylation and RNP assembly by immunoprecipitation. Surprisingly, these studies revealed t hat the Sm-analogous region is not required either for binding of the commo n proteins or for cap trimethylation, Furthermore, except for the first 24 nt which are part of the U2 promoter, the U2 coding region could be substit uted or deleted without affecting cap trimethylation.