A Comparison of Cell Wall Proteomics between Clinical and Laboratory Saccharomyces cerevisiae Strains by Mass Spectrometric Analysis

• Main Authors: Pei-Chi Chiang, 江珮琪
• Other Authors: 張雅雯
• Format: Others
• Language: zh-TW
• Published: 2009
• Online Access: http://ndltd.ncl.edu.tw/handle/85040139863551564237

碩士 === 國立臺灣大學 === 醫學檢驗暨生物技術學研究所 === 97 === The yeast Saccharomyces cerevisiae is widely used in food industry and biotechnology. However, it has been regarded as an emerging fungal pathogen because of an increasing number of infections being reported in these two decades. In this study, we investigated the possible pathogenic mechanism of S. cerevisiae by characterizing the differences between the clinical and laboratory strains. First, we tried to set up an in vitro system of virulence determination, by which we can get some preliminary concepts of pathogenic potential of the yeast strains before in vivo murine model experiment. Previous study revealed that the virulence of S. cerevisiae strains correlated with the potential of proinflammtory cytokine induction in macrophages. We used the isolated peritoneal macrophages from BALA/c and C57BL/6 mice as the infected cells to do this test. However, we did not observe the correlation between TNF-α production levels and the pathogenicity of the clinical isolates we collected. The feasibility of the in vitro system should be further investigated. Fungal cells are surrounded by a cell wall, alterations of which are known to influence the host immune response after infection. MATH (Microbial Adhesion To Hydrocarbon) was used to characterize the cell surface property of different yeast strains, and the results showed that the clinical strains are more hydrophobic than the laboratory strains, indicating certain difference existed on the cell surface between these strains. The outermost part of yeast cell wall is composed of a dense layer of mannoproteins. To examine the difference in cell wall protein composition between the clinical and laboratory strains, we performed a mass spectrometry-based quantitative proteomics with the labeling strategy using SILAC (Stable Isotope Labeling by Amino acids in Cell culture). The expression levels of most cell wall proteins increased in the clinical strains we observed, especially for proteins Scw10p, Pst1p, and Hsp150p. Further study should be done to understand whether these proteins play important roles in the pathogenicity of the clinical strains. We propose that the clinical strains upregulate cell wall protein expression to form a dense coat, under which the potent proinflammatory β-glucan of inner cell wall cannot be recognize by host immune system. Thus, the yeast cells are capable of prolonged persistence in the host. However, this hypothesis needs further confirmation. Altogether, we reported here the characterization of the difference in cell wall proteomics that might be important to the pathogenicity of the clinical strains.