, 2 and 4 hours and total RNA was isolated followed by qRTPCR (See
, two and 4 hours and total RNA was isolated followed by qRTPCR (See Components and Solutions for specifics). As expected, expression of SFLTAP progressively improved from time points 0 h to 4 h (Figure 5A, left panel). This improved SFL expression correlated with decreased SFL2 and BRG expression (Figure 5A, middle 
and ideal Pyrroloquinolinequinone disodium salt web panels), constant having a unfavorable regulation of SFL2 andPLOS Pathogens plospathogens.orgBRG expression. On the other hand, PPCKinduced SFL2TAP expression (Figure 5B, left panel) correlated with decreased expression of SFL (Figure 5B, SFL panel) and enhanced expression of UME6 and ALS3 (Figure 5B, UME6 and ALS3 panels), constant with our microarray data (Figure four). Taken together, our transcriptomics information reflect the antagonistic functions of Sflp and Sfl2p in regulating C. albicans morphogenesis, with SFL promoting the yeastform development which correlates with upregulation of yeast formspecific genes and downregulation of HSGs, and SFL2 advertising hyphal development which correlates with upregulation of HSGs and downregulation of yeast formspecific genes.The Sflp and Sfl2p regulatory networkWe combined the transcriptomics as well as the ChIPSeq data so as to get a genomewide view on the transcriptional modules associated with Sflp and Sfl2p regulatory functions (Figure six). We have been expecting to PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/24121451 find a substantial amount of genes which are bound by Sflp and downregulated in pCaEXPSFLHA3 vs. pCaEXP microarray information, as Sflp is believed to act as a repressor. In line together with the function of Sfl2p as an activator of hyphal growth, we were also hypothesizing that binding of Sfl2p to its targets would correlate with improved expression of these target genes.C. albicans Sflp and Sfl2p Regulatory NetworksFigure six. Sflp and Sfl2p transcriptional modules. Venn diagrams of the overlap amongst the genes that happen to be modulated in (A) SFL or SFL2 transcriptomics (light red circles, upregulated; light green circles, downregulated; gene expression foldchange cutoff .5; Pvalue cutoff 0.05) and typically bound by Sflp and Sfl2p (light blue circle) or (B) SFL2 transcriptomics (light red circle, upregulated; light green circle, downregulated;PLOS Pathogens plospathogens.orgC. albicans Sflp and Sfl2p Regulatory Networksgene expression foldchange cutoff .5; Pvalue cutoff 0.05) and especially bound by Sfl2p (light grey circle). Numbers inside the Venn diagrams indicate the amount of genes. Circled numbers indicate the amount of genes that happen to be (A) each modulated in SFL or SFL2 transcriptomics data and frequently bound by Sflp and Sfl2p or (B) both modulated in SFL2 transcriptomics information and particularly bound by Sfl2p. The name of those genes (or their orf9 nomenclature) along with the functional categories to which they belong are shown inside the linked boxes. , DCK is essential for hyphal formation; orf9.3475 is a hyphal induced gene. doi:0.37journal.ppat.00359.gSurprisingly, amongst the 3 targets usually bound by Sflp and Sfl2p, 40 genes have been upregulated and only 22 genes had been downregulated in pCaEXPSFLHA3 vs. pCaEXP data (Figure 6A). Conversely, 39 genes have been downregulated in pCaEXPSFL2HA3 vs. pCaEXP information and only five genes were upregulated (Figure 6A), indicating that Sflp and Sfl2p have dual transcriptional regulatory functions; acting as each transcriptional activators and transcriptional repressors. As Sflp and Sfl2p respectively act as a repressor and an activator of hyphal development, we examined the set of genes that had been generally bound by these two regulators and whose exp.