Sjöstrand Jimmy O O, Kegel Andreas, Aström Stefan U
Developmental Biology, Wenner-Gren Institute, Arrhenius Laboratories E3, Stockholm University, SE-106 91 Stockholm, Sweden.
Eukaryot Cell. 2002 Aug;1(4):548-57. doi: 10.1128/EC.1.4.548-557.2002.
We studied the silencing of the cryptic mating-type loci HMLa and HMRa in the budding yeast Kluyveromyces lactis. A 102-bp minimal silencer fragment was defined that was both necessary and sufficient for silencing of HMLalpha. Mutagenesis of the silencer revealed three distinct regions (A, B, and C) that were important for silencing. Recombinant K. lactis ribosomal DNA enhancer binding protein 1 (Reb1p) could bind the silencer in vitro, and point mutations in the B box abolished both Reb1p binding and silencer function. Furthermore, strains carrying temperature-sensitive alleles of the REBI gene derepressed the transcription of the HMLalpha1 gene at the nonpermissive temperature. A functional silencer element from the K. lactis cryptic HMRa locus was also identified, which contained both Reb1p binding sites and A boxes, strongly suggesting a general role for these sequences in K lactis silencing. Our data indicate that different proteins bind to Kluyveromyces silencers than to Saccharomyces silencers. We suggest that the evolution of silencers is rapid in budding yeasts and discuss the similarities and differences between silencers in Saccharomyces and Kluyveromyces.
我们研究了乳酸克鲁维酵母中隐性交配型基因座HMLa和HMRa的沉默现象。确定了一个102碱基对的最小沉默子片段，它对于HMLα的沉默既必要又充分。对沉默子进行诱变揭示了三个对沉默很重要的不同区域（A、B和C）。重组的乳酸克鲁维酵母核糖体DNA增强子结合蛋白1（Reb1p）在体外能够结合沉默子，并且B框中的点突变消除了Reb1p的结合以及沉默子功能。此外，携带REBI基因温度敏感等位基因的菌株在非允许温度下会解除对HMLα1基因转录的抑制。还鉴定出了来自乳酸克鲁维酵母隐性HMRa基因座的一个功能性沉默子元件，它同时包含Reb1p结合位点和A框，这强烈表明这些序列在乳酸克鲁维酵母的沉默中具有普遍作用。我们的数据表明，与酿酒酵母的沉默子相比，不同的蛋白质与乳酸克鲁维酵母的沉默子结合。我们认为，出芽酵母中沉默子的进化很快，并讨论了酿酒酵母和乳酸克鲁维酵母中沉默子之间的异同。
