A temperature-responsive network links cell shape and virulence traits in a primary fungal pathogen -- RNAi mutants (2). Histoplasma capsulatum

The ability to grow at host temperature is a critical trait for most pathogenic microbes of humans. Thermally dimorphic fungal pathogens, including Histoplasma capsulatum, are a class of soil fungi that undergo a dramatic change in cell shape and virulence gene expression in response to host temperature. Here we elucidate a complex temperature-responsive network in H. capsulatum, which switches from an environmental filamentous form to a pathogenic yeast form. We dissect the circuit driven by three regulators that control yeast-phase growth, and demonstrate that these factors, including two deeply conserved Velvet family proteins of unknown function, associate with DNA. We identify and characterize a fourth regulator of this pathway, and define cis-acting motifs that recruit these transcription factors to a tightly interwoven network of temperature-responsive target genes. Our results provide the first comprehensive analysis of the complex transcriptional network that links temperature to morphology and virulence in thermally dimorphic fungi. This submission gives the expression profiling results. Overall design: cDNA from each ryp mutants and wild-type controls was labeled with Cy5 and competitively hybridized against the Cy3-labeled pooled reference sample using H. capsulatum whole-genome 70-mer oligonucleotide microarray. For the experiments with ryp T-DNA mutants, there were 4 to 6 replicates for each strain and condition, and for the experiments with ryp knockdown strains, there were 3 to 12 replicates for each strain and condition. The T-DNA and knockdown experiments are being submitted as separate series, with samples further divided based on G217B platform version.

在宿主体温下生长的能力，是大多数人类致病微生物的关键性状。温度双态性真菌致病菌（包括荚膜组织胞浆菌（Histoplasma capsulatum））属于一类土壤真菌，会响应宿主体温发生细胞形态与毒力基因表达的剧烈变化。本研究阐明了荚膜组织胞浆菌中一套复杂的温度响应调控网络——该菌可从环境丝状形态转换为致病酵母形态。我们解析了调控酵母相生长的三类调控因子所驱动的调控回路，并证实这些因子（包括两种功能未知的高度保守 Velvet家族蛋白（Velvet family proteins））可与DNA结合。我们还鉴定并表征了该通路中的第四种调控因子，并明确了可将这些转录因子招募至温度响应靶基因紧密互作网络中的顺式作用基序。本研究结果首次全面解析了温度双态性真菌中连接温度、形态与毒力的复杂转录调控网络。本次提交包含该表达谱分析的相关结果。实验整体设计：以Cy5标记每株ryp突变体与野生型对照的互补DNA（cDNA），并利用荚膜组织胞浆菌全基因组70-mer寡核苷酸微阵列，将其与Cy3标记的混合参考样本进行竞争性杂交。针对ryp T-DNA插入突变体的实验，每株菌株在各条件下均设置4至6次生物学重复；针对ryp基因敲低菌株的实验，每株菌株在各条件下则设置3至12次生物学重复。T-DNA插入突变与基因敲低实验将作为独立系列提交，样本还将根据G217B平台版本进一步分组。