Plasma membrane architecture protects Candida albicans from killing by copper

Abstract

The ability to resist copper toxicity is important for microbial pathogens to survive attack by innate immune cells. A sur7Δ mutant of the fungal pathogen Candida albicans exhibits decreased virulence that correlates with increased sensitivity to copper, as well as defects in other stress responses and morphogenesis. Previous studies indicated that copper kills sur7Δ cells by a mechanism distinct from the known resistance pathways involving the Crp1 copper exporter or the Cup1 metallothionein. Since Sur7 resides in punctate plasma membrane domains known as MCC/eisosomes, we examined overexpression of SUR7 and found that it rescued the copper sensitivity of a mutant that fails to form MCC/eisosomes (pil1Δ lsp1Δ), indicating that these domains act to facilitate Sur7 function. Genetic screening identified new copper-sensitive mutants, the strongest of which were similar to sur7Δ in having altered plasma membranes due to defects in membrane trafficking, cortical actin, and morphogenesis (rvs161Δ, rvs167Δ, and arp2Δ arp3Δ). Consistent with the mutants having altered plasma membrane organization, they were all more readily permeabilized by copper, which is known to bind phosphatidylserine and phosphatidylethanolamine and cause membrane damage. Although these phospholipids are normally localized to the intracellular leaflet of the plasma membrane, their exposure on the surface of the copper-sensitive mutants was indicated by increased susceptibility to membrane damaging agents that bind to these phospholipids. Increased copper sensitivity was also detected for a drs2Δ mutant, which lacks a phospholipid flippase that is involved in maintaining phospholipid asymmetry. Copper binds phosphatidylserine with very high affinity, and deleting CHO1 to prevent phosphatidylserine synthesis rescued the copper sensitivity of sur7Δ cells, confirming a major role for phosphatidylserine in copper sensitivity. These results highlight how proper plasma membrane architecture protects fungal pathogens from copper and attack by the immune system, thereby opening up new avenues for therapeutic intervention.

The Composition and the Structure of MCC/Eisosomes in Neurospora crassa

“…It raises the possibility MCC/eisosomes may have function(s) in protein modification and secretion. Furthermore, GlcN6P is an essential precursor of cell wall components chitin (Bulik et al, 2003) and NCU04639 also has functional annotations in the cell wall macromolecule metabolic process (GO-score = 0.44), which indicates that MCC/eisosomes, consistent with previous data, may be required for proper cell wall morphogenesis including Pil1, Lsp1, Sur7, and Slm1/2 (Alvarez et al, 2008;Wang et al, 2011Wang et al, , 2016Douglas et al, 2013;Douglas and Konopka, 2019). Additionally, NCU05230 and NCU04639 both have a transmembrane receptor functional annotation and the former is related to light stimulus detection (GOscore = 0.56) while the latter is associated with defense response (Cscores GO = 0.22, GO-score = 0.35).…”

Fungal plasma membrane domains

“…This is related to altered organization of the cell wall, as sur7Δ cells have thicker but weaker cell wall and show extensive, aberrant invaginations of the cell wall (Alvarez, Douglas and Konopka 2009;Wang et al 2011;Douglas et al 2012;Foderaro, Douglas and Konopka 2017). The same phenotypes are observed in pil1Δ lsp1Δ cells, and interestingly they can be rescued by overexpression of Sur7, suggesting that for these phenotypes eisosomes act by promoting the proper localization and/or stability of Sur7 (Wang et al 2011(Wang et al , 2016Douglas and Konopka 2019).…”

Copper blocks V‐ATPase activity and SNARE complex formation to inhibit yeast vacuole fusion

“…For example, in HeLa cells, 10 to 100 μM Cu 2+ is needed to activate the PI3 Kinase/AKT signaling pathway . Others have grown Candida albicans on 10 to 500 μM CuSO 4 to find mutants that affect resistance to copper . To show copper toxicity in growing Saccharomyces cerevisiae , Sowada et al used millimolar concentrations of CuCl 2 .…”

“…15 Others have grown Candida albicans on 10 to 500 μM CuSO 4 to find mutants that affect resistance to copper. 16 To show copper toxicity in growing Saccharomyces cerevisiae, Sowada et al 17 used millimolar concentrations of CuCl 2 . Finally, a recent proteomic and genetic analysis of the response to soluble copper by yeast used ≥200 μM CuSO 4 to screen their collection of strains.…”