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.

Reciprocal functions of Cryptococcus neoformans copper homeostasis machinery during pulmonary infection and meningoencephalitis

“…To survive, replicate and invade host tissues, pathogenic fungi have evolved efficient and sophisticated machineries to compete for essential nutrients, including metals. In the last decade, studies have implied that Cu is a key virulence modulator for C. neoformans and is required for fungal virulence 13,14,18,[26][27][28] . Fungal pathogens require Cu for maintaining normal cell growth and are equipped with Cu-detoxification defences, suggesting the possibility that fungal cells encounter both Cu scarcity and toxicity during their infectious life cycles.…”

“…29, revealed that a cuf1D strain is sensitive to both low and high Cu 12,29 . A subsequent study showed that Cuf1 controls the expression of the Cu transporter CTR4 and that the cuf1D strain is avirulent in a tail vein infection model, thus suggesting that Cu acquisition is essential for C. neoformans virulence in mice 18,27 . However, recent work has demonstrated that Cuf1 activates both the Cu-detoxification machinery and the Cu-acquisition machinery, in response to high or low Cu conditions, respectively 12 .…”

“…For example, two independent studies demonstrated robust cell growth of cuf1D strains under nutrient-deficient condition 12,32 , given Cuf1 is a positive regulator of CTR4 (ref. 12,27). The nutrient phenotype of the ctr4D strain has not been reproduced while generating the ctr4D mutant using other markers, such as NEO, HYG or URA5 (Supplementary Table, Figs 1 and 4, Supplementary Figs 2 and 3) 12 .…”

“…Ctr4 has been hypothesized to be important for cryptococcal meningoencephalitis 18,27 . Our results provide the first evidence that C. neoformans switches from Cu detoxification to acquisition modes when it disseminates from the lung to the brain (Fig.…”

Contribution of ATPase copper transporters in animal but not plant virulence of the crossover pathogen Aspergillus flavus

“…In the human pathogen Cryptococcus neoformans , MTs play a major role of in Cu detoxification [7]. Two MTs, Cmt1 and Cmt2, were shown to be important for fungal virulence in C. neoformans , both of which are regulated by the Cu-binding TF Cuf1 [2,18]. In Candida albicans , a membrane-localized P-type Cu ATPase, Crp1, is responsible for Cu resistance along with the MT Cup1, both of which are transcriptionally activated by Ace1 under high Cu conditions [19–21].…”