Characterization of a candidate Trypanosoma brucei U1 small nuclear RNA gene

We have previously shown that the poly(A) polymerase (PAP) gene of Trypanos oma brucei is interrupted by an intervening sequence. It was postulated tha t removing this intron by cis-splicing requires a yet unidentified U1 small nuclear RNA (snRNA), which in other organisms engages in base-pair interac tions across the 5 ' splice site during early spliceosome assembly. Here we present a characterization of a 75 nucleotide long candidate T. brucei U1 snRNA. Immunoprecipitation studies indicate that a trimethylguanosine cap s tructure is present at the 5 ' end and that the RNA is: bound to core prote ins common to spliceosomal ribonucleoprotein particles. The U1 snRNA has th e potential for extensive intermolecular base pairing with the PAP 5 ' spli ce site. We used block replacement mutagenesis to identify sequences necess ary for in vivo expression of U1 snRNA. We found that at least two cis-acti ng elements, tRNA-like A and B boxes, located in the 5 ' -flanking region a re necessary for U1 snRNA synthesis; no internal sequences close to the tra nscription start site are essential, suggesting a promoter architecture dis tinct from other trypanosome U-snRNA genes. (C) 2001 Elsevier Science B.V. All rights reserved.