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Small-Scale Plasma Membrane Preparation for the Analysis of Candida albicans Cdr1-mGFPHis
Published on: June 13, 2021
Eisosomes promote the ability of Sur7 to regulate plasma membrane organization in Candida albicans
Hong X Wang1, Lois M Douglas1, Petra Veselá2
1Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, NY 11794-5222.
Abstract:
The plasma membrane of the fungal pathogen Candida albicans forms a protective barrier that also mediates many processes needed for virulence, including cell wall synthesis, invasive hyphal morphogenesis, and nutrient uptake. Because compartmentalization of the plasma membrane is believed to coordinate these diverse activities, we examined plasma membrane microdomains termed eisosomes or membrane compartment of Can1 (MCC), which correspond to ∼200-nm-long furrows in the plasma membrane. A pil1∆ lsp1∆ mutant failed to form eisosomes and displayed strong defects in plasma membrane organization and morphogenesis, including extensive cell wall invaginations. Mutation of eisosome proteins Slm2, Pkh2, and Pkh3 did not cause similar cell wall defects, although pkh2∆ cells formed chains of furrows and pkh3∆ cells formed wider furrows, identifying novel roles for the Pkh protein kinases in regulating furrows. In contrast, the sur7∆ mutant formed cell wall invaginations similar to those for the pil1∆ lsp1∆ mutant even though it could form eisosomes and furrows. A PH-domain probe revealed that the regulatory lipid phosphatidylinositol 4,5-bisphosphate was enriched at sites of cell wall invaginations in both the sur7∆ and pil1∆ lsp1∆ cells, indicating that this contributes to the defects. The sur7∆ and pil1∆ lsp1∆ mutants displayed differential susceptibility to various types of stress, indicating that they affect overlapping but distinct functions. In support of this, many mutant phenotypes of the pil1∆ lsp1∆ cells were rescued by overexpressing SUR7 These results demonstrate that C. albicans eisosomes promote the ability of Sur7 to regulate plasma membrane organization.
Insights
Candida albicans plasma membrane organization is crucial for virulence. Eisosomes, plasma membrane microdomains, are vital for regulating cell wall synthesis and morphogenesis, with Sur7 playing a key role.
Area of Science:
- Mycology
- Cell Biology
- Biochemistry
Background:
- The plasma membrane of Candida albicans is essential for virulence, mediating processes like cell wall synthesis and morphogenesis.
- Plasma membrane compartmentalization into microdomains, such as eisosomes (membrane compartment of Can1 or MCC), is thought to coordinate these functions.
- Eisosomes are characterized as approximately 200-nm-long furrows within the plasma membrane.
Purpose of the Study:
- To investigate the role of eisosomes and associated proteins in Candida albicans plasma membrane organization and morphogenesis.
- To identify specific proteins and lipids involved in eisosome function and their contribution to virulence-related processes.
- To elucidate the relationship between eisosomes, the Sur7 protein, and plasma membrane integrity.
Main Methods:
- Genetic manipulation of eisosome components (pil1∆ lsp1∆, slm2∆, pkh2∆, pkh3∆, sur7∆ mutants).
- Microscopic analysis of plasma membrane organization, cell wall integrity, and furrow formation.
- Lipid localization studies using PH-domain probes to detect phosphatidylinositol 4,5-bisphosphate enrichment.
Main Results:
- The pil1∆ lsp1∆ mutant lacked eisosomes and exhibited significant defects in plasma membrane organization and morphogenesis, including cell wall invaginations.
- While mutations in Slm2, Pkh2, and Pkh3 did not cause similar cell wall defects, Pkh2 and Pkh3 kinases were implicated in furrow regulation.
- The sur7∆ mutant displayed cell wall invaginations similar to the pil1∆ lsp1∆ mutant, and phosphatidylinositol 4,5-bisphosphate was enriched at invagination sites in both mutants.
- sur7∆ and pil1∆ lsp1∆ mutants showed differential stress susceptibility, and overexpressing SUR7 rescued many pil1∆ lsp1∆ phenotypes.
Conclusions:
- C. albicans eisosomes are critical for proper plasma membrane organization and morphogenesis.
- The Sur7 protein, in conjunction with eisosomes, plays a significant role in regulating plasma membrane organization.
- Dysregulation of eisosome function and Sur7 leads to cell wall defects and altered stress responses, impacting fungal virulence.
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