Role of the ESCRT Pathway in Laccase Trafficking and Virulence of Cryptococcus neoformans

Abstract

The endosomal sorting complex required for transport (ESCRT) plays a crucial role in the transportation and degradation of proteins. We determined that Vps27, a key protein of the ESCRT-0 complex, is required for the transport of the virulence factor laccase to the cell wall in Cryptococcus neoformans. Laccase activity was perturbed, as was melanin production, in vps27Δ strains. In the absence of VPS27, there was an accumulation of multivesicular bodies with vacuolar fragmentation and mistargeting of the vacuolar carboxypeptidase CPY/Prc1, resulting in an extracellular localization. In addition, deletion of VPS27 resulted in a defect in laccase targeting of a Lac1-green fluorescent protein (GFP) fusion to the cell wall with trapping within intracellular puncta; this deletion was accompanied by reduced virulence in a mouse model. However, the actin cytoskeleton remained intact, suggesting that the trafficking defect is not due to defects in actin-related localization. Extracellular vesicle maturation was also defective in the vps27Δ mutant, which had a larger vesicle size as measured by dynamic light scattering. Our data identify cryptococcal VPS27 as a required gene for laccase trafficking and attenuates virulence of C. neoformans in a mouse intravenous (i.v.) meningitis model.

Internalization of the host alkaline pH signal in a fungal pathogen

“…CME is initiated by the recruitment of coat proteins and clathrin to membrane-bound receptor-ligand complexes that are targeted for internalization. These coated regions of the membrane invaginate to form endocytic vesicles, which are then transported to intracellular micro-niches including the Endosomal Sorting Complex Required for Transport (ESCRT) (Conner and Schmid, 2003; Gonzá Lez-Gaitá N and Stenmark, 2003; Hurley and Emr, 2006; Miaczynska and Stenmark, 2008; Park et al ., 2020). Likely due to their involvement in the transport of internalized cellular material, ESCRT proteins are required for stress tolerance, including the adaptation of microbial pathogens to extracellular conditions encountered in the infected human host ([8,23,41–43].…”

“…These coated regions of the membrane invaginate to form endocytic vesicles, which are then transported to intracellular micro-niches including the Endosomal Sorting Complex Required for Transport (ESCRT) (Conner and Schmid, 2003; Gonzá Lez-Gaitá N and Stenmark, 2003; Hurley and Emr, 2006; Miaczynska and Stenmark, 2008; Park et al ., 2020). Likely due to their involvement in the transport of internalized cellular material, ESCRT proteins are required for stress tolerance, including the adaptation of microbial pathogens to extracellular conditions encountered in the infected human host ([8,23,41–43]. In many fungi such as the human pathogens C. albicans, A fumigatus , and C. neoformans , and the plant pathogen Rhizoctonia solani , this endocytosis process is required for growth and differentiation in response to changes in the extracellular environment [8,23,35,43–45].…”

“…Likely due to their involvement in the transport of internalized cellular material, ESCRT proteins are required for stress tolerance, including the adaptation of microbial pathogens to extracellular conditions encountered in the infected human host ([8,23,41–43]. In many fungi such as the human pathogens C. albicans, A fumigatus , and C. neoformans , and the plant pathogen Rhizoctonia solani , this endocytosis process is required for growth and differentiation in response to changes in the extracellular environment [8,23,35,43–45]. Accordingly, disrupting protein trafficking pathways often results in defective fungal virulence [23,43].…”

“…In many fungi such as the human pathogens C. albicans, A fumigatus , and C. neoformans , and the plant pathogen Rhizoctonia solani , this endocytosis process is required for growth and differentiation in response to changes in the extracellular environment [8,23,35,43–45]. Accordingly, disrupting protein trafficking pathways often results in defective fungal virulence [23,43].…”

Cross-Kingdom Extracellular Vesicles EV-RNA Communication as a Mechanism for Host–Pathogen Interaction

“…The deletion of Sec6, a component of the exocyst complex involved with vesicles fusion with the plasma membrane, prevents EVs production and laccase secretion to extracellular milieu, and decreased virulence in mice (Panepinto et al, 2009). In addition, mutations in protein of the ESCRT complex, Vps27, led to MVB accumulation and decreased laccase transport to the cell wall (Park et al, 2020).…”