Conservation and Evolution of Cis-Regulatory Systems in Ascomycete Fungi

Relatively little is known about the mechanisms through which gene expression regulation evolves. To investigate this, we systematically explored the conservation of regulatory networks in fungi by examining the cis-regulatory elements that govern the expression of coregulated genes. We first identified groups of coregulated Saccharomyces cerevisiae genes enriched for genes with known upstream or downstream cis-regulatory sequences. Reasoning that many of these gene groups are coregulated in related species as well, we performed similar analyses on orthologs of coregulated S. cerevisiae genes in 13 other ascomycete species. We find that many species-specific gene groups are enriched for the same flanking regulatory sequences as those found in the orthologous gene groups from S. cerevisiae, indicating that those regulatory systems have been conserved in multiple ascomycete species. In addition to these clear cases of regulatory conservation, we find examples of cis-element evolution that suggest multiple modes of regulatory diversification, including alterations in transcription factor-binding specificity, incorporation of new gene targets into an existing regulatory system, and cooption of regulatory systems to control a different set of genes. We investigated one example in greater detail by measuring the in vitro activity of the S. cerevisiae transcription factor Rpn4p and its orthologs from Candida albicans and Neurospora crassa. Our results suggest that the DNA binding specificity of these proteins has coevolved with the sequences found upstream of the Rpn4p target genes and suggest that Rpn4p has a different function in N. crassa.目前学界对基因表达调控的演化机制尚缺乏深入认知。为探究该问题，我们通过分析调控共调控基因表达的顺式调控元件（cis-regulatory elements），系统探究了真菌调控网络的保守性。我们首先鉴定出多组共调控的酿酒酵母（Saccharomyces cerevisiae）基因簇，这些基因簇中富集了携带已知上游或下游顺式调控序列的基因。鉴于这些基因簇在近缘物种中大概率同样存在共调控现象，我们对另外13种子囊真菌物种中酿酒酵母共调控基因的直系同源基因（orthologs）开展了类似分析。我们发现，诸多物种特异性基因簇所富集的侧翼调控序列，与酿酒酵母同源基因簇中的侧翼调控序列一致，这表明这些调控系统在多种子囊真菌中得以保守保留。除上述明确的调控保守案例外，我们还观测到顺式元件演化的实例，其揭示了多种调控多样化模式：包括转录因子结合特异性的改变、将新基因靶标整合至现有调控系统，以及共招募调控系统以调控另一组基因。我们选取其中一个实例展开深入研究：检测了酿酒酵母转录因子Rpn4p，以及白色念珠菌（Candida albicans）和粗糙脉孢霉（Neurospora crassa）中其同源蛋白的体外活性。我们的研究结果表明，这些蛋白的DNA结合特异性与Rpn4p靶基因上游序列发生了协同演化，同时也揭示出Rpn4p在粗糙脉孢霉中行使着截然不同的功能。