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J. Biol. Chem., Vol. 283, Issue 21, 14327-14334, May 23, 2008

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Cryptococcal Xylosyltransferase 1 (Cxt1p) from Cryptococcus neoformans Plays a Direct Role in the Synthesis of Capsule Polysaccharides^*

J. Stacey Klutts and Tamara L. Doering¹

From the Departments of Molecular Microbiology and Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri 63110

The opportunistic yeast Cryptococcus neoformans causes serious disease in humans and expresses a prominent polysaccharide capsule that is required for its virulence. Little is known about how this capsule is synthesized. We previously identified a β1,2-xylosyltransferase (Cxt1p) with in vitro enzymatic activity appropriate for involvement in capsule synthesis. Here, we investigate C. neoformans strains in which the corresponding gene has been deleted (cxt1). Loss of CXT1 does not affect in vitro growth of the mutant cells or the general morphology of their capsules. However, NMR structural analysis of the two main capsule polysaccharides, glucuronoxylomannan (GXM) and galactoxylomannan (GalXM), showed that both were missing β1,2-xylose residues. There was an 30% reduction in the abundance of this residue in GXM in mutant compared with wild-type strains, and mutant GalXM was almost completely devoid of β1,2-linked xylose. The GalXM from the mutant strain was also missing a β1,3-linked xylose residue. Furthermore, deletion of CXT1 led to attenuation of cryptococcal growth in a mouse model of infection, suggesting that the affected xylose residues are important for normal host-pathogen interactions. Cxt1p is the first glycosyltransferase with a defined role in C. neoformans capsule biosynthesis, and cxt1 is the only strain identified to date with structural alterations of the capsule polysaccharide GalXM.

Received for publication, October 30, 2007 , and in revised form, March 14, 2008.

^* This work was supported, in whole or in part, by National Institutes of Health Grants GM R01 071007 (to T. L. D.) and GM F32 072341 (to J. S. K.). This work was also supported by Department of Energy Center for Plant and Microbial Complex Carbohydrates Grant DE-FG09-93ER-20097 and a William Keck Foundation Postdoctoral Fellowship (to J. S. K.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Tables S1–S3 and Figs. S1–S3.

¹ To whom correspondence should be addressed: 660 South Euclid Ave., Campus Box 8230, St. Louis, MO, 63110-1093. Tel.: 314-747-5597; Fax: 314-362-1232; E-mail: doering{at}wustl.edu.

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