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Extension of O-linked mannosylation in the Golgi is critical for cell wall integrity signaling and interaction with host cells in Cryptococcus neoformans pathogenesis

ABSTRACT: Purpose: To identify the downstream gene sets regulated by Mpk1p in C. neoformans normal and Tunicamycin-treated conditions, we compared the RNA-seq based transcriptional profiles. Methods: Total RNA was isolated according to the RNeasy Mini Kit (Qiagen) and all procedure for RNA sequencing were conducted by ebiogen. The mRNA was isolated by using the Poly(A) RNA Selection Kit (LEXOGEN) and isolated mRNAs were used for the cDNA synthesis. Results: Transcriptome analysis of the wild-type strain revealed that tunicamycin significantly increased expression of a subset of calcineurin-Crz1 pathway related genes , but most of which were not upregulated in O-glycan mutant strain. Conclusion: Our results support the essential role of O-glycan in maintaining cell wall integrity upon stress conditions of C. neoformans.

ORGANISM(S): Cryptococcus neoformans

PROVIDER: GSE198875 | GEO | 2022/11/11

REPOSITORIES: GEO

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Project description:Calcineurin is a highly conserved Ca2+/calmodulin-dependent serine/threonine-specific protein phosphatase that orchestrates cellular Ca2+ signaling responses. In Cryptococcus neoformans, calcineurin is activated by multiple stresses including high temperature, and is essential for stress adaptation and virulence. The transcription factor Crz1 is a major calcineurin effector in Saccharomyces cerevisiae and other fungi. Calcineurin dephosphorylates Crz1, thereby enabling Crz1 nuclear translocation and transcription of target genes. Here we show that Crz1 is dephosphorylated by calcineurin, and crz1Δ mutants display phenotypes intermediate between wild-type and calcineurin mutants. RNA-sequencing revealed 102 genes that are regulated in a calcineurin/Crz1-dependent manner at 37°C. 99 genes were down-regulated in cna1Δ and crz1Δ mutants, indicating these genes are normally activated by the calcineurin/Crz1 pathway at high temperature. About 58% of calcineurin-Crz1 target genes have unknown functions, while genes with known or predicted functions are involved in cell wall remodeling, calcium transport, and pheromone production. Surprisingly, only five genes have orthologs known to be calcineurin/Crz1-dependent in S. cerevisiae. 393 genes are independently regulated by calcineurin, and Crz1 regulates 59 genes independently of calcineurin. Taken together, these results indicate that the calcineurin/Crz1-dependent pathway controls a transcriptional circuit that has been extensively rewired in C. neoformans compared to S. cerevisiae. Given the intermediate crz1Δ mutant phenotype, and our recent evidence for a calcineurin regulatory network impacting mRNA in P-bodies and stress granules independently of Crz1, calcineurin likely acts on factors beyond Crz1 that govern mRNA expression/stability to operate a branched transcriptional/post-transcriptional stress response network necessary for fungal virulence. Taken together, our findings reveal the core calcineurin-Crz1 stress response cascade is maintained from ascomycetes to a pathogenic basidiomycete fungus, but its output has been extensively rewired to promote fungal virulence.

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Extension of O-linked mannosylation in the Golgi is critical for cell wall integrity signaling and interaction with host cells in Cryptococcus neoformans pathogenesis

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