Gene expression in human fungal pathogen Coccidioides immitis changes as arthroconidia differentiate into spherules and mature

Abstract

BackgroundCoccidioides immitis is a dimorphic fungus that causes disease in mammals, including human beings. It grows as a mycelium containing arthroconidia in the soil and in the host arthroconidia differentiates into a unique structure called a spherule. We used a custom open reading frame oligonucleotide microarray to compare the transcriptome of C. immitis mycelia with early (day 2) and late stage (day 8) spherules grown in vitro. All hybridizations were done in quadruplicate and stringent criteria were used to identify significantly differentially expressed genes.Results22% of C. immitis genes were differentially expressed in either day 2 or day 8 spherules compared to mycelia, and about 12% of genes were differentially expressed comparing the two spherule time points. Oxireductases, including an extracellular superoxide dismutase, were upregulated in spherules and they may be important for defense against oxidative stress. Many signal transduction molecules, including pleckstrin domain proteins, protein kinases and transcription factors were downregulated in day 2 spherules. Several genes involved in sulfur metabolism were downregulated in day 8 spherules compared to day 2 spherules. Transcription of amylase and α (1,3) glucan synthase was upregulated in spherules; these genes have been found to be important for differentiation to yeast in Histoplasma. There were two homologs of 4-hydroxyphenylpyruvate dioxygenase (4-HPPD); transcription of one was up- and the other downregulated. We tested the effect of a 4-HPPD inhibitor, nitisinone, on mycelial and spherule growth and found that it inhibited mycelial but not spherule growth.ConclusionsTranscription of many genes was differentially expressed in the process of arthroconidia to spherule conversion and spherule maturation, as would be expected given the magnitude of the morphologic change. The transcription profile of early stage (day 2) spherules was different than late stage (day 8) endosporulating spherules. In addition, very few genes that are important for spore to yeast conversion in other dimorphic fungi are differentially expressed in C. immitis mycelia and spherules suggesting that dimorphic fungi may have evolved different mechanisms to differentiate from mycelia to tissue invasive forms.

Coccidioides immitis and posadasii; A review of their biology, genomics, pathogenesis, and host immunity

“…There are only two published studies comparing the transcriptome of mycelia to that of spherules grown in vitro; one studied both C. immitis and C. posadasii using RNA-seq and the other studied only C. immitis , using a whole genome open reading frame microarray [41,42]. The RNA-seq study compared mycelia and day 4 spherules and concentrated on genes that were differentially expressed in both species.…”

“…Oxireductases (including extracellular superoxide dismutase that could help protect against neutrophil killing) are up-regulated in day 2 spherules, as are sugar transporters, thioesterase, and amylase. About a third of genes that are up-regulated in vivo have no assigned function and 5% are only found in Coccidioides spp [42]. One of the most interesting gene families that are down-regulated in C. immitis spherules is the protein kinase family; 28 of 184 predicted protein kinase genes were downregulated.…”

“…Venn diagram of C. immitis genes up-regulated more than 2-fold compared to mycelial gene expression. (a) The data for day 4 spherules are from [41] and (b) the data for Day 2 and day 8 spherules are from [42]. …”

“…Carbohydrate metabolic pathways found in the 152 genes up-regulated in day 2, 4 and 8 spherules [41,42]. a – all genes with that designation found in the genome; b – genes with that designation found in the 152 genes identified.…”

The Quest for a Vaccine against Coccidioidomycosis, A Neglected Disease of the Americas

“…A single spherule can contain hundreds of endospores ( Figure 2B). There have been three studies of the global gene expression in mycelia and spherules by RNAseq and microarray [16][17][18]. Between 7-20% of genes were differentially expressed in spherules compared to mycelia.…”