Studies on the Roles of Clathrin-Mediated Membrane Trafficking and Zinc Transporter Cis4 in the Transport of GPI-Anchored Proteins in Fission Yeast

Wurentuya Jaiseng,Yan Ma,Takayoshi Kuno,Yue Fang,Reiko Sugiura,Christopher Beh

doi:10.1371/journal.pone.0041946

Wurentuya Jaiseng, Yan Ma + Show 4 more

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https://doi.org/10.1371/journal.pone.0041946

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Abstract

We previously identified Cis4, a zinc transporter belonging to the cation diffusion facilitator protein family, and we demonstrated that Cis4 is implicated in Golgi membrane trafficking in fission yeast. Here, we identified three glycosylphosphatidylinositol (GPI)-anchored proteins, namely Ecm33, Aah3, and Gaz2, as multicopy suppressors of the MgCl2-sensitive phenotype of cis4-1 mutant. The phenotypes of ecm33, aah3 and gaz2 deletion cells were distinct from each other, and Cis4 overexpression suppressed Δecm33 phenotypes but did not suppress Δaah3 defects. Notably, green fluorescent protein-tagged Ecm33, which was observed at the cell surface in wild-type cells, mostly localized as intracellular dots that are presumed to be the Golgi and endosomes in membrane-trafficking mutants, including Δapm1, ypt3-i5, and chc1-1 mutants. Interestingly, all these membrane-trafficking mutants showed hypersensitivity to BE49385A, an inhibitor of Its8 that is involved in GPI-anchored protein synthesis. Taken together, these results suggest that GPI-anchored proteins are transported through a clathrin-mediated post-Golgi membrane trafficking pathway and that zinc transporter Cis4 may play roles in membrane trafficking of GPI-anchored proteins in fission yeast.

Highlights

Glycosylphosphatidylinositol (GPI) anchoring is a common post-translational lipid modification by which proteins are attached to the cell surface in all eukaryotic cells
In another screen using FK506, we identified a mutant allele of the cis4+ gene that encodes a zinc transporter belonging to the cation diffusion facilitator (CDF) protein family, and we characterized the role of Cis4 in Golgi membrane trafficking in fission yeast [11]
We have previously demonstrated that Cis4 is a zinc transporter belonging to the CDF protein family, and plays a role in Golgi membrane trafficking in fission yeast [11]

Summary

Introduction

Glycosylphosphatidylinositol (GPI) anchoring is a common post-translational lipid modification by which proteins are attached to the cell surface in all eukaryotic cells. GPI-anchored proteins are functionally diverse and are important for signal transduction, cell-cell interaction, cell adhesion, cell surface protection, and cell wall synthesis [1,2,3,4]. In budding yeast Saccharomyces cerevisiae, more than 60 genes are predicted to encode GPI-anchored proteins that play important roles in cell wall biogenesis and cell wall assembly [7,8]. In the fission yeast Schizosaccharomyces pombe, 33 GPI-anchored protein candidates have been identified among 4950 S. pombe ORFs [9]. We identified a mutation in the its8+ gene encoding a homolog of the budding yeast Mcd4p and human Pig-n that are involved in GPI anchor synthesis through a genetic screen using the immunosuppressant drug FK506, a specific inhibitor of calcineurin [10]

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