Cell surface mannan oligosaccharides have been implicated in die pathogenesis of C. albicans through roles such as adhesion to host cells and modulation of the immune system (Calderone, 1993, Chaffin, 1998). Understanding the molecular biology of glycosylation is therefore critical to elucidating host-fungus interactions. The CaMNTI gene, responsible for adding the second mannose on 0-linked mannan has previously been shown to be important for adhesion and virulence in C. albicans (Buurman et ai, 1998). In this project, the CaMNT1 gene family was studied as a means to understand more about the roles of oligosaccharide synthesis in Candida-host interactions. The Camntlp enzyme was expressed heterologously in P. pastoris and its cofactor and acceptor specificities were determined. The key residues for the retaining mechanism of CaMntlp catalysis and for cofactor binding were determined by site-directed mutagenesis as Asp350, Glu-318, His-377 and His377. This knowledge may aid in the rational design of antifungals against the MNT1 family of Candida mannosyltransferases. Two further members of the gene fainily; CaMNT3 and CaMNT5 were disrupted using the ura-blaster technique. Deletion of CaMNT3 and to a lesser extent CaMNT5 led to strains unable to form hyphae on solid Spider medium. This phenotype is distinct from that reported for CaMNT1 and suggests a role for these genes upstream or downstream of the Cphlp MAP Itinase cascade that regulates yeast-hypha morphogenesis. The Camnt3 and CamntS nuU mutants were attenuated in virulence in a systemic mouse model of candidosis. The heterozygous mutants were more attenuated in virulence than the nuU mutants indicating that a compensation mechanism may upregulate expression of other MNT genes when both copies of CaMNTS or CaMNT3 are disrupted. This study has shown that glycan synthesis affects morphogenesis in C. albicans and indicates many cell wall proteins require to be glycosylated to perform their function.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:323140 |
| Date | January 2000 |
| Creators | Thomson, Lynn |
| Publisher | University of Aberdeen |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://digitool.abdn.ac.uk/R?func=search-advanced-go&find_code1=WSN&request1=AAIU602055 |
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