1887

Abstract

The gene encodes a unique 1,3-β-glucosyltransferase (BgI2p) present in the cell wall of and other fungi. Although believed to be involved in cell wall assembly, disruption of the gene in showed no apparent phenotype. We performed sequential disruptions of the loci in a homozygous clinical isolate of using the blaster method, in order to investigate the role of BgI2p in this dimorphic, pathogenic fungus. Strain CACW-1 contained disruptions of both homologues of the gene and lacked BgI2p, as assessed by protein extraction, SDS-PAGE and Western blot analysis, and enzyme assay; however, residual non-BgI2p transferase activity was detected. CACW-1 was attenuated in virulence for mice when compared to an isogenic parent strain, and fewer organisms were recovered from the kidneys of infected animals. Additional phenotypic changes included: (1) a dramatic increase in the sensitivity to the chitin synthesis inhibitor nikkomycin Z when CACW-1 cells were incubated at 37 or 42 °; (2) an 8.7+1.6% slower growth rate at 37 ° for CACW-1 when compared to its isogenic parent; and (3) aggregation of CACW-1 cells during stationary phase and/or incubation of stationary phase cells in phosphate buffer. Characterization of SDS-extracted cell walls did not reveal any significant differences in the levels of 1,3-β- or 1,6-β-glucan. These data reveal that loss of BgI2p does have a phenotype in and indicate that (1) loss of BgI2p function renders cells more dependent on chitin for wall integrity, and attenuates virulence (probably due to subtle changes in wall structure), and (2) that additional 1,3-β-glucosyltransferases are present in the disruptant.

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1997-02-01
2021-05-06
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