Expression of Penicillium marneffei chitin synthase genes in response to cell-wall stressors

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Expression of Penicillium marneffei chitin synthase genes in response to cell-wall stressors

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Title: Expression of Penicillium marneffei chitin synthase genes in response to cell-wall stressors
Author: Engle, Joshua
Abstract: Penicillium marneffei is a dimorphic fungus and an opportunistic pathogen responsible for the disease penicilliosis. Efforts to genetically characterize this organism have led to the development of a plasmid-dependent, mutant generating system known as Agrobacterium tumefaciens mediated transformation. This system produced one mutant, originally identified as strain I231, which was found to harbor a defective yakA gene. As determined by the work of Suwunnakorn et al. (2014), this defective gene resulted in an abnormal phenotype and unique stress responses when compared to a wild-type strain. The yakA mutant possessed increased resistance to the anionic dye Congo Red (CR), known to inhibit glucan synthase activity, yet also yielded increased sensitivity to the similarly acting antifungal drug, caspofungin. These seemingly contradictory results, along with a increase in overall chitin content within the mutant, led to this study which seeks to determine overall chitin synthase gene expression in the yakA mutant, the wild-type strain, and the complement strain, CY21, in terms of seven known chitin synthase genes. These strains were exposed to CR, the detergent sodium dodecyl sulfate (SDS), the chitin binding agent Calcofluor White (CW), as well as incubated under non-stressed control conditions. Subsequently, RNA from these cultures were isolated and chitin synthase gene expression levels were quantitated through qRT-PCR using the reference gene benA (encoding ß-tubulin) for normalization. The results of this study indicated that the interrupted yakA gene produced no significant effect in terms of chitin synthase gene expression under stressed conditions, but the mutation produced significantly higher expression levels of the chitin synthases in the yakA control conditions at both 25 C and 37 C. This response most likely represents a compensatory response to the weakened cell wall caused by an interrupted yakA gene.
Description: xi, 86 leaves : illustrations ; 29 cm
URI: http://hdl.handle.net/1989/11705
Date: 2015

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