Expression of peptidase genes of the isolate HM-1:IMSS as determined by microarrays

<p><b>Copyright information:</b></p><p>Taken from "The genome: primary structure and expression of proteolytic enzymes"</p><p>http://www.biomedcentral.com/1471-2164/8/170</p><p>BMC Genomics 2007;8():170-170.</p><p>Published online 14 Jun 2007</p><p>PMCID:PMC1913524.</p><p></p> Error bars represent the standard error of the mean of nine hybridizations (biological replicates)

SomA interacts with PtaB in <i>A</i>. <i>fumigatus</i>.

<p>(A) The abundance of SomA and PtaB was measured by LC/MS and estimated based on MaxQuant’s logarithmized label free quantification (log2 LFQ) intensities. High and intermediate LFQ intensities are shown in red and blue. Absence of peptides and low LFQ intensities are presented in black. (B) Western hybridization of GFP-Trap and RFP-Trap enrichments with α-GFP antibody. The single band in the RFP-Trap indicated by an arrow was identified as SomA-GFP by LC/MS with the given peptides. (C) Reciprocal western to (B) but an α-RFP antibody instead of the α-GFP antibody as a different probe. The double band (arrow) correspond to PtaB and SomA by LC/MS. For all experiments, the strains were grown in MM medium for 24 h at 37°C. Protein extracts were performed with either GFP-Trap or RFP-Trap beads and the eluted proteins were separated by 12% SDS-PAGE.</p

Transcription Factor SomA Is Required for Adhesion, Development and Virulence of the Human Pathogen <i>Aspergillus fumigatus</i>

<div><p>The transcription factor Flo8/Som1 controls filamentous growth in <i>Saccharomyces cerevisiae</i> and virulence in the plant pathogen <i>Magnaporthe oryzae</i>. Flo8/Som1 includes a characteristic N-terminal LUG/LUH-Flo8-single-stranded DNA binding (LUFS) domain and is activated by the cAMP dependent protein kinase A signaling pathway. Heterologous SomA from <i>Aspergillus fumigatus</i> rescued in yeast <i>flo8</i> mutant strains several phenotypes including adhesion or flocculation in haploids and pseudohyphal growth in diploids, respectively. <i>A</i>. <i>fumigatus</i> SomA acts similarly to yeast Flo8 on the promoter of <i>FLO11</i> fused with reporter gene (<i>LacZ</i>) in <i>S</i>. <i>cerevisiae</i>. <i>FLO11</i> expression in yeast requires an activator complex including Flo8 and Mfg1. Furthermore, SomA physically interacts with PtaB, which is related to yeast Mfg1. Loss of the <i>somA</i> gene in <i>A</i>. <i>fumigatus</i> resulted in a slow growth phenotype and a block in asexual development. Only aerial hyphae without further differentiation could be formed. The deletion phenotype was verified by a conditional expression of <i>somA</i> using the inducible Tet-on system. A adherence assay with the conditional <i>somA</i> expression strain indicated that SomA is required for biofilm formation. A <i>ptaB</i> deletion strain showed a similar phenotype supporting that the SomA/PtaB complex controls <i>A</i>. <i>fumigatus</i> biofilm formation. Transcriptional analysis showed that SomA regulates expression of genes for several transcription factors which control conidiation or adhesion of <i>A</i>. <i>fumigatus</i>. Infection assays with fertilized chicken eggs as well as with mice revealed that SomA is required for pathogenicity. These data corroborate a complex control function of SomA acting as a central factor of the transcriptional network, which connects adhesion, spore formation and virulence in the opportunistic human pathogen <i>A</i>. <i>fumigatus</i>.</p></div

Model of the SomA and SomA-PtaB genetic network in <i>A</i>. <i>fumigatus</i>.

<p>The solid arrows indicate the presented data and the results of previous studies [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005205#ppat.1005205.ref025" target="_blank">25</a>, <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005205#ppat.1005205.ref028" target="_blank">28</a>–<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005205#ppat.1005205.ref030" target="_blank">30</a>, <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005205#ppat.1005205.ref056" target="_blank">56</a>]. We showed that SomA/PtaB complex is a transcriptional activator for downstream targets (<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005205#ppat.1005205.g006" target="_blank">Fig 6</a> and <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005205#ppat.1005205.s003" target="_blank">S3 Fig</a>) and these proteins had different cellular functions in <i>A</i>. <i>fumigatus</i> (See <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005205#sec010" target="_blank">Discussion</a>).</p

SomA interacts with PtaB in <i>A</i>. <i>fumigatus</i>.

<p>(A) The abundance of SomA and PtaB was measured by LC/MS and estimated based on MaxQuant’s logarithmized label free quantification (log2 LFQ) intensities. High and intermediate LFQ intensities are shown in red and blue. Absence of peptides and low LFQ intensities are presented in black. (B) Western hybridization of GFP-Trap and RFP-Trap enrichments with α-GFP antibody. The single band in the RFP-Trap indicated by an arrow was identified as SomA-GFP by LC/MS with the given peptides. (C) Reciprocal western to (B) but an α-RFP antibody instead of the α-GFP antibody as a different probe. The double band (arrow) correspond to PtaB and SomA by LC/MS. For all experiments, the strains were grown in MM medium for 24 h at 37°C. Protein extracts were performed with either GFP-Trap or RFP-Trap beads and the eluted proteins were separated by 12% SDS-PAGE.</p

SomA regulates genes for conidiation and adhesion.

<p>Relative expression of genes encoding proteins that regulate conidiation and adhesion in the <i>Tet-somA</i> strain. The <i>Tet-somA</i> strain was cultivated in liquid MM medium for 18 h at 37°C and shifted to (A) liquid MM medium for 8 h at 37°C and (B) solid MM plate for 24 h at 37°C. Addition of 5 mg/L doxycycline is indicated as (<i>On</i>). Levels for the <i>Tet-somA On</i> state were set to 1. Graph indicates mean ± standard errors from two independent experiments.</p

SomA is required for virulence in an egg and a mouse model of invasive aspergillosis.

<p>(A) Survival rate of eggs inoculated with the indicated strains. Addition of doxycycline was not performed in the <i>somA</i> complemented mutant and <i>Tet-somA Off</i> state. (B) Survival rate of mice infected with the indicated strains. (C) Histological pictures of mouse lungs obtained from indicated dates after infection with the corresponding strains or PBS.</p

Fungal strains used in this study.

<p>Fungal strains used in this study.</p

SomA promotes growth and conidia formation of <i>A</i>. <i>fumigatus</i>.

<p>(A) Colony morphology and growth rate of the indicated strains. All strains were grown on either MM plate or MM plate with 5 mg/L doxycycline indicated as (+) for 5 days at 37°C. Values in the graph are indicated as means ± standard error. (B) Morphology of conidiation in the indicated strains. Upper panel: Strains were grown on MM or MM with 5 mg/L doxycycline agar-coated slides for 28 h at 37°C. The open arrows indicate conidiophores and filled arrows represent the vesicles for sporulation. Lower panel: Strains were grown on MM or MM with 5 mg/L doxycycline agar-slides for 28 h at 37°C. Scale bars represent 20 μm (upper panel) and 50 μm (lower panel). Strains grow on MM agar with doxycycline indicated as (+). For all experiments, the <i>Tet-somA</i> was induced (<i>On</i> state) at the present of doxycycline.</p

Plasmids used in this study.

<p>Plasmids used in this study.</p