Failure to detect binding of Trypanosoma brucei SNAPc to U2 and U6 snRNA gene sequences by in vitro transcription competition and pull-down assays.
The small nuclear RNA (snRNA)-activating protein complex (SNAP(c)) is a multi-subunit transcription factor characterized in humans and Drosophila melanogaster. It binds to an upstream sequence element (USE) of snRNA gene promoters and activates both RNA polymerase (pol) II and III-mediated transcription of snRNA genes. The first identified and partially characterized transcription factor in a trypanosomatid organism appears to be a SNAP(c) homologue. It was identified in Leptomonas seymouri and shown to specifically interact with the USE of the RNA pol II-transcribed spliced leader (SL) RNA gene. Recently, chromatin immunoprecipitation and a gel shift assay suggested that L. seymouri SNAP(c) also interacts with RNA pol III-transcribed U2 and U6 snRNA genes. Previously, we have characterized and epitope-tagged the Trypanosoma brucei homologue of the SNAP50 subunit. Here, we show by in vitro transcription competition and promoter pull-down assays that TbSNAP50 binds to the SL RNA gene promoter and parasite-specifically to the ribosomal RNA gene promoter. Conversely, we did not detect binding of the factor to U2 and U6 snRNA gene sequences. Since U snRNA gene promoters are structurally conserved among trypanosomatids, our findings contrast those obtained in L. seymouri and suggest that trypanosomatid SNAP(c) is not involved in RNA pol III-mediated transcription of U snRNA genes.
Schimanski
Bernd
B
Center for Microbial Pathogenesis, University of Connecticut Health Center, 263 Farmington Avenue, Farmington, CT 06030-3710, USA.
Ma
Luyan
L
Günzl
Arthur
A
D000818:Animals; D004268:DNA-Binding Proteins / Q000378:metabolism*; D017125:Genes, Protozoan; D006801:Humans; D066298:In Vitro Techniques; D015800:Protozoan Proteins / Q000378:metabolism*; D016053:RNA, Protozoan / Q000235:genetics* / Q000378:metabolism*; D012342:RNA, Small Nuclear / Q000235:genetics* / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D014346:Trypanosoma brucei brucei / Q000235:genetics* / Q000378:metabolism*
D004268:DNA-Binding Proteins; D015800:Protozoan Proteins; D016053:RNA, Protozoan; D012342:RNA, Small Nuclear; D014157:Transcription Factors; C405484:U2 small nuclear RNA; C405486:U6 small nuclear RNA