Biebinger S, Rettenmaier S, Flaspohler J, Hartmann C, Peña-Diaz J, Wirtz L E, Hotz H R, Barry J D, Clayton C
Zentrum für Molekulare Biologie, Universität Heidelberg, Germany.
Nucleic Acids Res. 1996 Apr 1;24(7):1202-11. doi: 10.1093/nar/24.7.1202.
African trypanosomes are extracellular protozoan parasites that are transmitted from one mammalian host to the next by tsetse flies. Bloodstream forms express variant surface glycoprotein (VSG); the tsetse fly (procyclic) forms express instead the procyclic acidic repetitive protein (PARP). PARP mRNA is abundant in procyclic forms and almost undetectable in blood-stream forms. Post-transcriptional mechanisms are mainly responsible for PARP mRNA regulation but results of nuclear run-on experiments suggested that transcription might also be regulated. We measured the activity of genomically-integrated PARP, VSG and rRNA promoters in permanently-transformed blood-stream and procyclic form trypanosomes, using reporter gene constructs that showed no post-transcriptional regulation. When the constructs were integrated in the rRNA non-transcribed spacer, the ribosomal RNA and VSG promoters were not developmentally regulated, but integration at the PARP locus reduced rRNA promoter activity in bloodstream forms. PARP promoter activity was 5-fold down-regulated in bloodstream forms when integrated at either site. Regulation was probably at the level of transcriptional initiation, but elongation through plasmid vector sequences was also reduced.
非洲锥虫是细胞外原生动物寄生虫,通过采采蝇从一个哺乳动物宿主传播到另一个宿主。血流型表达可变表面糖蛋白(VSG);采采蝇(前循环型)则表达前循环酸性重复蛋白(PARP)。PARP mRNA在前循环型中丰富,在血流型中几乎检测不到。转录后机制主要负责PARP mRNA的调控,但核转录实验结果表明转录也可能受到调控。我们使用无转录后调控的报告基因构建体,测量了基因组整合的PARP、VSG和rRNA启动子在永久转化的血流型和前循环型锥虫中的活性。当构建体整合到rRNA非转录间隔区时,核糖体RNA和VSG启动子不受发育调控,但整合到PARP基因座会降低血流型中rRNA启动子的活性。当整合到任一位置时,PARP启动子活性在血流型中下调5倍。调控可能发生在转录起始水平,但通过质粒载体序列的延伸也减少了。