The calcium transporter Pmc1 provides Ca²⁺ tolerance and influences the progression of murine cryptococcal infection

Abstract

The Ca2+‐calcineurin signaling pathway in the human fungal pathogen Cryptococcus neoformans is essential for adaptation to the host environment during infection. Calcium transporters regulate cytosolic calcium concentrations, providing Ca2+ loading into storage organelles. The three calcium transporters that have been characterized in C. neoformans, Cch1, Eca1 and Vcx1, are required for fungal virulence, supporting a role for calcium‐mediated signaling in cryptococcal pathogenesis. In the present study, we report the functional characterization of the putative vacuolar calcium ATPase Pmc1 in C. neoformans. Our results demonstrate that Pmc1 provides tolerance to high Ca2+ concentrations. The double knockout of C. neoformans PMC1 and VCX1 genes impaired the intracellular calcium transport, resulting in a significant increase in cytosolic calcium levels. Furthermore, Pmc1 was essential for both the progression of pulmonary infection and brain colonization in mice, emphasizing the crucial role of calcium signaling and transport for cryptococcal pathogenesis.

Microevolution of Serial Clinical Isolates ofCryptococcus neoformansvar.grubiiandC. gattii

“…CDA2 (CNAG_01230) and CDA3 (CNAG_01239) encode two of the three deacetylases that account for the production of chitosan, which is necessary for maintenance of the cryptococcal cell wall and capsule (34). PMC1 (CNAG_01232) encodes a calcium transporter, which provides Ca 2+ tolerance and is essential for the progression of pulmonary infection in mice and dissemination to the brain (35). HapX (CNAG_01242) regulates iron acquisition and metabolism (36), and a recent study of the other iron acquisition gene, FRE3 , showed that this type of gene can be viewed as a “virulence adaptation gene” and evolved in mammalian hosts (37).…”

Role of the Apt1 Protein in Polysaccharide Secretion by Cryptococcus neoformans

“…However, a number of secretory mutants of C. neoformans showed normal capsular dimensions but reduced concentrations of extracellular GXM (35,51,58,59). This information might suggest that the efficiency of C. neoformans in synthesizing and exporting GXM may exceed the minimum quantitative requirements for full capsule formation.…”

“…In this study, we demonstrated a previously unknown function of flippases in physiological and pathogenic secretion-related events used by C. neoformans. To the best of our knowledge, cryptococcal regulators of secretion described so far include the products of the SEC (51,58,64) and CAP genes (65-70), Golgi reassembly and stacking protein (30), protein kinase A (71), vacuolar Ca 2ϩ transporters (35,59), and the vacuolar protein Vps23 (72). Mutants with altered expression of the genes coding for each of these secretory regulators are hypovirulent or avirulent in mice.…”

Components of the Calcium-Calcineurin Signaling Pathway in Fungal Cells and Their Potential as Antifungal Targets

“…The concentrations of cytosolic Ca 2ϩ are very low at resting states, ranging from 50 to 200 nM in fungal cells when the environmental Ca 2ϩ concentrations range from Ͻ1 M to Ͼ100 mM (26,28,29). The calcium homeostasis system, which consists of various calcium channels and pumps, as well as many related proteins and enzymes, plays an important role in maintaining the optimal Ca 2ϩ concentrations in the cytosol and intracellular compartments, such as the vacuole, endoplasmic reticulum (ER), and Golgi apparatus (29)(30)(31). In general, the plasma membrane Ca 2ϩ influx system is activated to result in a rapid influx of Ca 2ϩ ions in response to various external stresses, such as store-operated stress, hyperosmotic stress, alkaline stress, cold stress, thermal stress, oxidative stress, and ethanol stress (32)(33)(34)(35)(36)(37).…”

“…Numerous enzymes that catalyze the folding, modification, processing, and trafficking of secretory proteins are activated in response to stress, which results in Ca 2ϩ sequestration in secretory organelles or the initiation of other restorative pathways. Specifically, the Ca 2ϩ -ATPase Pmc1p and H ϩ /Ca 2ϩ exchanger Vcx1p, which are located on the vacuole membrane, and the Ca 2ϩ pumps Pmr1p, Cod1p, and Eca1p, which are predominantly located on the Golgi apparatus and ER, are activated to direct the cytosolic Ca 2ϩ to secretory organelles, such as the vacuole, Golgi apparatus, and ER (31,58,(62)(63)(64)(65). However, Vcx1p was identified as the protein complex that is predominantly responsible for restoring cytosolic Ca 2ϩ concentrations after a brief challenge with high extracellular Ca 2ϩ concentrations, while Pmc1p appears to be critical for long-term Ca 2ϩ tolerance (59).…”