Characterizing the role and function of two hybrid histidine kinases in histoplasma capsulatum

Daniel W. Grangaard, The College of Wooster

Abstract

Histoplasma capsulatum is a dimorphic fungal pathogen that exists in two distinct and different morphologies: mold and yeast. H. capsulatum has the ability to infect a wide range and variety of mammalian hosts, including most commonly humans and household pets. A host becomes infected from H. capsulatum by inhaling the mold phase spores (microcondia), which once inside the host undergo a rapid morphological transition from the mold phase to the yeast phase. It has been previously demonstrated that this morphological change, while required for virulence within a host, can be induced strictly by an increase in temperature from 25°C to 37°C. Specific signal transduction proteins, hybrid histidine kinases (HHK), have been implicated in not only sensing but also in mediating the necessary cellular response, via changes in gene expression, to allow the organism to survive. These HHK proteins have been identified in a multitude of dimorphic fungal pathogens including H. capsulatum. The aim of this study was to identify two specific HHK in H. capsulatum and examine their cellular role in mediating the gene expression required for the survival of H. capsulatum in a variety of environmental conditions. The approach to this experimentation was to block the expression of these two HHK genes through the means of RNA interference (RNAi). Based on evidence supporting the importance of HHK in their ability to mediate the necessary gene expression, it was hypothesized that the RNAi mutants created should not be able to sustain sufficient cell growth in an array of environmental conditions. In this study, two hybrid histidine kinase genes, HcHHK1 and HcHHK2 were identified in H. capsulatum, but were ultimately not able to be examined in any experimentation using 2 RNAi. While the components required for constructing the RNAi inducing vector were collected for the HcHHK1 gene, they were never successfully recombined together.