Role of a CUF1/CTR4 copper regulatory axis in the virulence of Cryptococcus neoformans
Scott R. Waterman, Moshe Hacham, Guowu Hu, Xudong Zhu, Yoon-Dong Park, Soowan Shin, John Panepinto, Tibor Valyi-Nagy, Craig Beam, Shahid Husain, Nina Singh, Peter R. Williamson
Scott R. Waterman, Moshe Hacham, Guowu Hu, Xudong Zhu, Yoon-Dong Park, Soowan Shin, John Panepinto, Tibor Valyi-Nagy, Craig Beam, Shahid Husain, Nina Singh, Peter R. Williamson
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Research Article Microbiology

Role of a CUF1/CTR4 copper regulatory axis in the virulence of Cryptococcus neoformans

Abstract

The study of regulatory networks in human pathogens such as Cryptococcus neoformans provides insights into host-pathogen interactions that may allow for correlation of gene expression patterns with clinical outcomes. In the present study, deletion of the cryptococcal copper-dependent transcription factor 1 (Cuf1) led to defects in growth and virulence factor expression in low copper conditions. In mouse models, cuf1Δ strains exhibited reduced dissemination to the brain, but no change in lung growth, suggesting copper is limiting in neurologic infections. To examine this further, a biologic probe of available copper was constructed using the cryptococcal CUF1-dependent copper transporter, CTR4. Fungal cells demonstrated high CTR4 expression levels after phagocytosis by macrophage-like J774.16 cells and during infection of mouse brains, but not lungs, consistent with limited copper availability during neurologic infection. This was extended to human brain infections by demonstrating CTR4 expression during C. neoformans infection of an AIDS patient. Moreover, high CTR4 expression by cryptococcal strains from 24 solid organ transplant patients was associated with dissemination to the CNS. Our results suggest that copper acquisition plays a central role in fungal pathogenesis during neurologic infection and that measurement of stable traits such as CTR4 expression may be useful for risk stratification of individuals with cryptococcosis.

Authors

Scott R. Waterman, Moshe Hacham, Guowu Hu, Xudong Zhu, Yoon-Dong Park, Soowan Shin, John Panepinto, Tibor Valyi-Nagy, Craig Beam, Shahid Husain, Nina Singh, Peter R. Williamson

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Figure 1

Deletion of CUF1 from C. neoformans yields a copper-sensitive phenotype.

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Deletion of CUF1 from C. neoformans yields a copper-sensitive phenotype....
(A) Schematic of the Cuf1 protein from C. neoformans shows a KGRP consensus sequence and multiple cysteine-rich domains (cysteines represented by circles). (B) Phenotypes of CUF1 mutants under various copper conditions. Cells were diluted to an A600 of 0.2, serially diluted 1:5, and applied to YPD alone or YPD supplemented with either 750 μM BCS or 10 μM CuCl2 at 37°C for 3 days. (C) Cells were incubated for melanin production as described in Methods (left panels) or on malt extract for 5 days at 30°C and examined by India ink (middle panels) or incubated on Christensen’s agar for 2 hours (right panels).