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Understanding the internalization dynamics of N‐acetylglucosamine transporter (Ngt1) in Candida albicans
Author(s) -
Rao Kongara Hanumantha
Publication year - 2019
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2019.33.1_supplement.465.2
Subject(s) - internalization , endocytosis , microbiology and biotechnology , transporter , glucose transporter , biology , ubiquitin , candida albicans , lipid raft , cell membrane , chemistry , biochemistry , cell , signal transduction , gene , insulin , endocrinology
Integral membrane proteins are subjected to tight regulation in response to environmental cues. Ubiquitylation mediated endocytosis forms an integral part of this machinery. However, such a dynamics has not been worked out for the novel Nacetylglucosamine transporter (Ngt1) in the pathogenic yeast Candida albicans . N‐acetylglucosamine (GlcNAc) stimulates a wide range of signalling events in several organisms apart from evoking the phenomenon of virulence in several pathogens. Therefore, immense importance lies in the proper understanding of its uptake before the initiation of signalling. In this study we have investigated the Ngt1 Post Translational Modifications (PTMs) and its interaction with membrane lipids during internalization on cell membrane. We employed western blot, FACS, lipid binding assay, LC‐MS/MS and mutant analysis to understand the various key events involved in internalization. Our studies demonstrate that there exists sophisticated and subtle phosphorylation dependent ubiquitylation machinery that is responsible for proper maintenance of the transporter level at the cell surface. Further our study highlights how the transporter interacts with components of membrane microdomains (lipid rafts) like ergosterol for modulation of the transporter activity. Being a robust transporter, it can provide a tool for development of an inducible promoter and can help in designing therapeutic reagents to modulate GlcNAc entry. Support or Funding Information Council of Scientific and Industrial Research (CSIR), IndiaLocalization dynamics of GlcNA transporter (Ngt1) upon glucose addition At 90 min of GlcNAc induction almost all the fraction of Ngt1‐GFP resides in the cell membrane. Immediately within 5 min after addition of glucose some fraction protein which internalized into cytosol appeared as dotted punctate like structures (Early‐endosomes, late‐endosomes, MultiVesicularBodies, TransGolgi) and within 30–60 min, the degraded fraction appeared in vacuole, merged with vacuolar dye, CMAC‐CellTracker Blue.Life Cycle of Ngt1 Ngt1 is a membrane resident protein having 12 membrane spanning domains as predicted by TMHMM server v. 2.0 Ngt1 senses very minute concentrations of sugar. It undergoes sugar‐responsive ubiquitylation followed by its degradation within the vacuole. This provides the pathogen a better way to respond to their environment and simultaneously influence surface remodelling at the plasma membrane. The transporter shows association with novel components of the lipid rafts.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .