Polarized Exocytosis Induces Compensatory Endocytosis by Sec4p-Regulated Cortical Actin Polymerization

Cytokinesis is a complicated yet conserved step of the cell-division cycle that requires the coordination of multiple proteins and cellular processes. Here we describe a previously uncharacterized protein, Ync13, and its roles during fission yeast cytokinesis. Ync13 is a member of the UNC-13/Munc13 protein family, whose animal homologues are essential priming factors for soluble N-ethylmaleimide-sensitive factor attachment protein receptor complex assembly during exocytosis in various cell types, but no roles in cytokinesis have been reported. We find that Ync13 binds to lipids in vitro and dynamically localizes to the plasma membrane at cell tips during interphase and at the division site during cytokinesis. Deletion of Ync13 leads to defective septation and exocytosis, uneven distribution of cell-wall enzymes and components of cell-wall integrity pathway along the division site and massive cell lysis during cell separation. Interestingly, loss of Ync13 compromises endocytic site selection at the division plane. Collectively, we find that Ync13 has a novel function as an UNC-13/Munc13 protein in coordinating exocytosis, endocytosis, and cell-wall integrity during fission yeast cytokinesis.

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“…The balance between exocytosis and endocytosis is critical for plasma membrane dynamics ( Johansen et al. , 2016 ; Xie et al.…”

Section: Resultsmentioning

confidence: 99%

“…, 2005 ). In budding yeast, the polarized exocytosis and endocytosis are coupled by Rab GTPase Sec4 ( Johansen et al. , 2016 ).…”

Section: Discussionmentioning

confidence: 99%

Roles of the fission yeast UNC-13/Munc13 protein Ync13 in late stages of cytokinesis

Zhu

Hyun

Pan

et al. 2018

MBoC

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“…Confocal fluorescence microscopy was performed as previously described [ 93 ]. For all experiments, yeast cells were grown to mid-log phase before visualization.…”

Section: Methodsmentioning

confidence: 99%

Endoplasmic reticulum-plasma membrane contact sites integrate sterol and phospholipid regulation

et al. 2018

Self Cite

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Tether proteins attach the endoplasmic reticulum (ER) to other cellular membranes, thereby creating contact sites that are proposed to form platforms for regulating lipid homeostasis and facilitating non-vesicular lipid exchange. Sterols are synthesized in the ER and transported by non-vesicular mechanisms to the plasma membrane (PM), where they represent almost half of all PM lipids and contribute critically to the barrier function of the PM. To determine whether contact sites are important for both sterol exchange between the ER and PM and intermembrane regulation of lipid metabolism, we generated Δ-super-tether (Δ-s-tether) yeast cells that lack six previously identified tethering proteins (yeast extended synatotagmin [E-Syt], vesicle-associated membrane protein [VAMP]-associated protein [VAP], and TMEM16-anoctamin homologues) as well as the presumptive tether Ice2. Despite the lack of ER-PM contacts in these cells, ER-PM sterol exchange is robust, indicating that the sterol transport machinery is either absent from or not uniquely located at contact sites. Unexpectedly, we found that the transport of exogenously supplied sterol to the ER occurs more slowly in Δ-s-tether cells than in wild-type (WT) cells. We pinpointed this defect to changes in sterol organization and transbilayer movement within the PM bilayer caused by phospholipid dysregulation, evinced by changes in the abundance and organization of PM lipids. Indeed, deletion of either OSH4, which encodes a sterol/phosphatidylinositol-4-phosphate (PI4P) exchange protein, or SAC1, which encodes a PI4P phosphatase, caused synthetic lethality in Δ-s-tether cells due to disruptions in redundant PI4P and phospholipid regulatory pathways. The growth defect of Δ-s-tether cells was rescued with an artificial "ER-PM staple," a tether assembled from unrelated non-yeast protein domains, indicating that endogenous tether proteins have nonspecific bridging functions. Finally, we discovered that sterols play a role in regulating ER-PM contact site formation. In sterol-depleted cells, levels of the yeast E-Syt tether Tcb3 were induced and ER-PM contact increased dramatically. These results support a model in which ER-PM contact sites provide a nexus for coordinating the complex interrelationship between sterols, sphingolipids, and phospholipids that maintain PM composition and integrity.

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“…albicans as a human fungal pathogen results in part from its ability to switch between different morphological states, in order to adapt to the diverse challenges of the human host [ 12 – 15 ]. The spatio-temporal regulation of exocytosis and endocytosis is likely to be crucial for growth and morphology changes [ 16 , 17 ], and these two processes are critical for hyphal growth (reviewed in [ 18 ]). For example in C .…”

Section: Introductionmentioning

confidence: 99%

Role of Arf GTPases in fungal morphogenesis and virulence

Labbaoui¹,

Bogliolo²,

Ghugtyal³

et al. 2017

PLoS Pathog

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Virulence of the human fungal pathogen Candida albicans depends on the switch from budding to filamentous growth, which requires sustained membrane traffic and polarized growth. In many organisms, small GTPases of the Arf (ADP-ribosylation factor) family regulate membrane/protein trafficking, yet little is known about their role in fungal filamentous growth. To investigate these GTPases in C. albicans, we generated loss of function mutants in all 3 Arf proteins, Arf1-Arf3, and 2 Arf-like proteins, Arl1 and Arl3. Our results indicate that of these proteins, Arf2 is required for viability and sensitivity to antifungal drugs. Repressible ARF2 expression results in defects in filamentous growth, cell wall integrity and virulence, likely due to alteration of the Golgi. Arl1 is also required for invasive filamentous growth and, although arl1/arl1 cells can initiate hyphal growth, hyphae are substantially shorter than that of the wild-type, due to the inability of this mutant to maintain hyphal growth at a single site. We show that this defect does not result from an alteration of phospholipid distribution and is unlikely to result from the sole Golgin Imh1 mislocalization, as Imh1 is not required for invasive filamentous growth. Rather, our results suggest that the arl1/arl1 hyphal growth defect results from increased secretion in this mutant. Strikingly, the arl1/arl1 mutant is drastically reduced in virulence during oropharyngeal candidiasis. Together, our results highlight the importance of Arl1 and Arf2 as key regulators of hyphal growth and virulence in C. albicans and identify a unique function of Arl1 in secretion.

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Polarized Exocytosis Induces Compensatory Endocytosis by Sec4p-Regulated Cortical Actin Polymerization

Cited by 25 publications

References 79 publications

Roles of the fission yeast UNC-13/Munc13 protein Ync13 in late stages of cytokinesis

Roles of the fission yeast UNC-13/Munc13 protein Ync13 in late stages of cytokinesis

Endoplasmic reticulum-plasma membrane contact sites integrate sterol and phospholipid regulation

Role of Arf GTPases in fungal morphogenesis and virulence

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