Table 2 qRT-PCR primers GAPDH S: ATTGGGCGTATTGTCTTCC AS: TTGAGCATGTAGGCAGCATA MAT1-1-1 S: TTCGTTCATAGCCTTCAGAAGCTTC AS: GGCCAGCATGACTGTCACGAAT PPG1 S: CTGTGTGGGCAGTAGCAATTCC AS: CAACGTTTCGCGACGAATTCA STE2 S: TGCTCTCGTTACTCGCAGAC AS: ATTGGATTATTGAGAAAATGGCTGGAATC AZD8931 order STE3 S: ACAATCGGTATATAACCAATACACAGTAG AS: GTTGTCCAGCACCGTCGATA BEM1 S: TGGAAGAAGATGACGGCGGAAT AS: TGTGGCTTTGTTGTAGGTGAGGG HMK1 S: CGTGGCAGCACAGACAATGC AS: GGCGGATTTGCAAGGACGT PKC1 S: CCGAAAGTCGTCACCAAGTG AS: CATGTAAGACTGCATTCTGAGC Western Blot An amount of organism equivalent to 10

μL was taken from an HMM plate kept at 37°C. 30 μL of Laemelli sample buffer was added and samples were boiled for 15 min. Samples were electrophoresed by SDS-PAGE using a precast 8-16% tris-glycine gel (Invitrogen) and transferred to a nitrocellulose membrane. Membranes were either incubated at room temperature for 2 hours with a 1:1000 dilution of anti-Myc alkaline phosphatase tagged antibody (Invitrogen), or with a 1:5000 dilution of rabbit anti-HSP60 antibody (a kind gift from Francisco Gomez, University of Cincinnati, Cincinnati, OH) as a loading control. Anti-HSP60 antibodies were secondarily tagged with a 1:1000 dilution of peroxidase labelled goat-anti-rabbit antibody (Kirkegaard and Perry Laboratories).

Phosphatase labelled antibodies Dinaciclib nmr were developed using BCIP/NBT phosphatase substrate (Kirkegaard and Perry Laboratories), and peroxidase labelled antibodies were developed using TMB One Component, HRP membrane substrate (BioFx Laboratories). Developed membranes were imaged using a FOTO/Analyst® FX system from Fotodyne®, Inc. Microarray Microarray analysis was performed in conjunction with the University of Cincinnati Danusertib Genomics and Microarray Thalidomide laboratory. Three samples of G217B and UC26 were grown at

25°C on nylon membranes placed on HMM plates as described above. RNA was extracted using TRIzol® reagent (Invitrogen) according to manufacturer’s instructions. Cy-3 and Cy-5 labelled cDNA from UC26 and G217B was hybridized to a slide containing 70-mer oligonucleotides representing each putative open reading frame in the H. capsulatum genome (Washington University Genome Sequencing Center, Washington University, St. Louis, MO). Dye swaps were also performed. Slides were imaged using a GenePixPro 4000 scanner (Axon Instruments), using Axon GenePix® Pro version 5.0 software. Cy3 and Cy5 intensities were normalized by subtracting local background intensities from the median intensity of each channel. Statistical analysis was performed by the University of Cincinnati Bioinformatics F&S Core of the Center for Environmental Genetics, as previously described [41]. Functional analysis was performed using BLAST2GO http://​www.​blast2go.