Regulatory networks that control fungal pathogenesis
Creators: name: University Of California Office Of The President
Publisher: name: University of California Office of the President
publisherIdentifier: https://ror.org/00dmfq477
publisherIdentifierScheme: ROR
Year: 2024

description: The goal of this project is to identify regulatory networks that control morphology and virulence in the systemic dimorphic fungal pathogens, which are important primary pathogens that infect mammalian hosts via inhalation of infectious particles produced in the soil. Our main focus is on the fungal pathogen Histoplasma capsulatum, which is thought to be responsible for approximately 500,000 infections per year in the U.S. alone. One of the most interesting aspects of H. capsulatum biology is that it changes its morphology and associated characteristics in response to its environment. In the soil, it grows in a hyphal (or filamentous) form. Inside the host, H. capsulatum undergoes a dramatic developmental change, switching its growth to yeast-form cells that colonize macrophages of the host immune system. The ability of cells to adopt this yeast-growth program is thought to be critical for H. capsulatum virulence. Furthermore, this morphologic switch can be recapitulated in culture by changing the temperature (room temperature growth triggers the soil form of the organism whereas growth at 37°C triggers the host form of the organism). My laboratory uses genetics and genomics to understand how temperature regulates morphology and virulence in this pathogen. We identified a transcription factor, Ryp1, which is a master transcriptional regulator required for yeast-phase growth. The ryp1 mutant is constitutively filamentous independent of temperature. Analysis of this mutant revealed that Ryp1 is required for regulation of the vast majority of genes that are enriched for expression at 37oC in wild-type cells. Here we propose to understand the regulatory circuits that are dependent on Ryp1 and are required for proper morphology and virulence at 37oC. With expert computational assistance from Patrik D'haeseleer at LLNL, in Aim 1 we will identify regulatory motifs that control gene expression by Ryp1 in response to temperature. Specifically, we have identified 50 genomic targets that associate with Ryp1 in H. capsulatum. We will use motif-finding algorithms to identify potential regulatory motifs in these target genes. Gel shift analyses and reporter assays will be used to determine whether these motifs are necessary and/or sufficient for association with Ryp1. In Aim 2, we will systematically disrupt the seven transcription factors that we have identified as direct downstream targets of Ryp1 and use whole genome expression profiling and computational analysis to determine the contribution of these individual transcription factors to the overall expression profile. Finally, we will utilize Dr. D'hesseleer's computational expertise to examine conservation of the resultant regulatory circuit in the systemic dimorphic fungi using recently available genome sequence for five different Histoplasma strains, as well as for Coccidioides, Blastomyces, and Paracoccidioides species. These comparisons will enable us to examine the evolution of this regulatory circuit among these human pathogens. The potential impact of these experiments is enormous. Even though the systemic dimorphic fungal pathogens are ubiquitous in the environment and adept at causing infection even in healthy hosts, very little is understood about how there pathogens adapt their morphogenetic program to effect host colonization in response to temperature. These studies represent the first exploration of temperature-dependent regulatory circuits in these organisms. If these studies identify a conserved regulatory circuit among the systemic dimorphic fungi, future experiments will directly address the function of this circuit in this group of pathogens as a whole.