The novel microtubule-associated CAP-glycine protein Cgp1 governs growth, differentiation, and virulence of Cryptococcus neoformans

Abstract

Microtubules are involved in mechanical support, cytoplasmic organization, and several cellular processes by interacting with diverse microtubule-associated proteins such as plus-end tracking proteins, motor proteins, and tubulin-folding cofactors. A number of the cytoskeleton-associated proteins (CAPs) contain the CAP-glycine-rich (CAP-Gly) domain, which is evolutionarily conserved and generally considered to bind to α-tubulin to regulate the function of microtubules. However, there has been a dearth of research on CAP-Gly proteins in fungal pathogens, including Cryptococcus neoformans, which is a global cause of fatal meningoencephalitis in immunocompromised patients. In this study, we identified five CAP-Gly protein-encoding genes in C. neoformans. Among these, Cgp1 encoded by CNAG_06352 has a unique domain structure containing CAP-Gly, SPEC, and Spc7 domains that is not orthologous to CAPs in other eukaryotes. Supporting the role of Cgp1 in microtubule-related function, we demonstrate that deletion or overexpression of CGP1 alters cellular susceptibility to thiabendazole, a microtubule destabilizer and that Cgp1 is co-localized with cytoplasmic microtubules. Related to the cellular function of microtubules, Cgp1 governs the maintenance of membrane stability and genotoxic stress responses. Deletion of CGP1 also reduces production of melanin pigment and attenuates the virulence of C. neoformans. Furthermore, we demonstrate that Cgp1 uniquely regulates the sexual differentiation of C. neoformans with distinct roles in the early and late stage of mating. Domain analysis revealed that the CAP-Gly domain plays a major role in all Cgp1 functions examined. In conclusion, this novel CAP-Gly protein, Cgp1, has pleotropic roles in regulating growth, stress responses, differentiation, and virulence in C. neoformans.

Fenbendazole controlsin vitrogrowth, virulence potential and animal infection in theCryptococcusmodel

“…Tubulin organization in C. neoformans and C. gattii was evaluated as described by Wang et al . (44) with minor modifications. Briefly, fungal cells were cultivated overnight in liquid Sabouraud medium at 30°C under shaking (200 rpm).…”

A unique cytoskeleton-associated protein in Cryptococcus neoformans

“…In a recent compelling study, Wang et al, identified a unique cytoskeleton-associated protein and examined its role in the pathogenesis of C. neoformans [ 5 ]. All eukaryotes consist of cytoskeletal filaments that include microtubules made from tubulin.…”

“…CAP-Gly proteins have been characterized in Saccharomyces cerevisiae and in Saccharomyces pombe , however the functional activity of CAP-Gly proteins in fungal pathogens are unknown thus underscoring the relevance of the study by Wang et al The goal of the study was to identify CAP-Gly proteins in Cryptococcus neoformans and to establish whether CAP-Gly proteins mediate mechanisms of pathogenesis [ 5 ]. The authors identified five genes predicted to encode CAP-Gly domains and subsequently used an in silico approach to analyze the protein domain structure of each gene.…”

“…Deletion of the Spc7 domain did not alter microtubule-related functions and sexual differentiation and the SPEC domain appeared to play only a very minor role in microtubule stability and filamentous growth. On the other hand, the CAP-Gly domain appeared to be required for all the functions observed for Cgp1 in C. neoformans [ 5 ].…”

“…Collectively, these multiple partners suggests that CAP-Gly proteins likely function in different biological processes. In fact, the study by Wang et al, found that Cgp1 appeared to have pleotropic roles in the biology and pathogenesis of C. neoformans [ 5 ].…”