Evidence of a role forS. cerevisiaeα-arrestin Art1 (Ldb19) in mating projection and zygote formations

When yeast cells sense mating pheromone, they undergo a characteristic response involving changes in transcription, cell cycle arrest in early G1, and polarization along the pheromone gradient. Cells in G2/M respond to pheromone at the transcriptional level but do not polarize or mate until G1. Fus2p, a key regulator of cell fusion, localizes to the tip of the mating projection during pheromone-induced G1 arrest. Although Fus2p was expressed in G2/M cells after pheromone induction, it accumulated in the nucleus until after cell division. As cells arrested in G1, Fus2p was exported from the nucleus and localized to the nascent tip. Phosphorylation of Fus2p by Fus3p was required for Fus2p export; cyclin/Cdc28p-dependent inhibition of Fus3p during late G1 through S phase was sufficient to block exit. However, during G2/M, when Fus3p was activated by pheromone signaling, Cdc28p activity again blocked Fus2p export. Our results indicate a novel mechanism by which pheromone-induced proteins are regulated during the transition from mitosis to conjugation.

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Dynamics of cell wall elasticity pattern shapes the cell during yeast mating morphogenesis

Open Biology ◽

10.1098/rsob.160136 ◽

2016 ◽

Vol 6 (9) ◽

pp. 160136 ◽

Cited By ~ 17

Author(s):

Björn Goldenbogen ◽

Wolfgang Giese ◽

Marie Hemmen ◽

Jannis Uhlendorf ◽

Andreas Herrmann ◽

...

Keyword(s):

Cell Wall ◽

Cell Shape ◽

Experimental Approach ◽

Time Resolved ◽

Force Microscopy ◽

Cell Wall Elasticity ◽

Atomic Force ◽

Stiff Material ◽

Mating Projection

The cell wall defines cell shape and maintains integrity of fungi and plants. When exposed to mating pheromone, Saccharomyces cerevisiae grows a mating projection and alters in morphology from spherical to shmoo form. Although structural and compositional alterations of the cell wall accompany shape transitions, their impact on cell wall elasticity is unknown. In a combined theoretical and experimental approach using finite-element modelling and atomic force microscopy (AFM), we investigated the influence of spatially and temporally varying material properties on mating morphogenesis. Time-resolved elasticity maps of shmooing yeast acquired with AFM in vivo revealed distinct patterns, with soft material at the emerging mating projection and stiff material at the tip. The observed cell wall softening in the protrusion region is necessary for the formation of the characteristic shmoo shape, and results in wider and longer mating projections. The approach is generally applicable to tip-growing fungi and plants cells.

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Coordinating cell polarization and morphogenesis through mechanical feedback

10.1101/2020.05.21.108076 ◽

2020 ◽

Cited By ~ 1

Author(s):

Samhita P. Banavar ◽

Michael Trogdon ◽

Brian Drawert ◽

Tau-Mu Yi ◽

Linda R. Petzold ◽

...

Keyword(s):

Cell Wall ◽

Cell Shape ◽

Cell Polarization ◽

Cell Wall Integrity ◽

Coarse Grained ◽

Cellular Processes ◽

Cell Wall Integrity Pathway ◽

Molecular Machinery ◽

Mating Projection ◽

Mechanical Feedback

AbstractMany cellular processes require cell polarization to be maintained as the cell changes shape, grows or moves. Without feedback mechanisms relaying information about cell shape to the polarity molecular machinery, the coordination between cell polarization and morphogenesis, movement or growth would not be possible. Here we theoretically and computationally study the role of a genetically-encoded mechanical feedback (in the Cell Wall Integrity Pathway) as a potential coordination mechanism between cell morphogenesis and polarity during budding yeast mating projection growth. We developed a coarse-grained continuum description of the coupled dynamics of cell polarization and morphogenesis as well as 3D stochastic simulations of the molecular polarization machinery in the evolving cell shape. Both theoretical approaches show that in the absence of mechanical feedback (or in the presence of weak feedback), cell polarity cannot be maintained at the projection tip during growth, with the polarization cap wandering off the projection tip, arresting morphogenesis. In contrast, for mechanical feedback strengths above a threshold, cells can robustly maintain cell polarization at the tip and simultaneously sustain mating projection growth. These results indicate that the mechanical feedback encoded in the Cell Wall Integrity pathway can provide important positional information to the molecular machinery in the cell, thereby enabling the coordination of cell polarization and morphogenesis.Author summaryCell migration, morphogenesis and secretion are among the vast number of cellular processes that require cells to define a preferred spatial direction to perform essential tasks. This is achieved by setting an intracellular molecular gradient that polarizes the cell. While the molecular players involved in cell polarization and some of the mechanisms that cells use to establish such molecular gradients are known, it remains unclear how cells maintain polarization as they dramatically change shape during morphogenesis, migration, etc. Here we identify a potential feedback control mechanism, encoded genetically in cells, that provides the molecular polarization machinery with the necessary information about cell geometry to maintain cell polarization during cell shape changes.

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Calcineurin, the Ca2+-dependent phosphatase, regulates Rga2, a Cdc42 GTPase-activating protein, to modulate pheromone signaling

Molecular Biology of the Cell ◽

10.1091/mbc.e16-06-0432 ◽

2017 ◽

Vol 28 (5) ◽

pp. 576-586 ◽

Cited By ~ 3

Author(s):

Nina Ly ◽

Martha S. Cyert

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Prolonged Exposure ◽

Receptor Internalization ◽

Gtpase Activating Protein ◽

Multiple Substrates ◽

Cycle Arrest ◽

Pheromone Signaling ◽

Mating Factor ◽

Mating Projection

Calcineurin, the conserved Ca2+/calmodulin-activated phosphatase, is required for viability during prolonged exposure to pheromone and acts through multiple substrates to down-regulate yeast pheromone signaling. Calcineurin regulates Dig2 and Rod1/Art4 to inhibit mating-induced gene expression and activate receptor internalization, respectively. Recent systematic approaches identified Rga2, a GTPase-activating protein (GAP) for the Cdc42 Rho-type GTPase, as a calcineurin substrate. Here we establish a physiological context for this regulation and show that calcineurin dephosphorylates and positively regulates Rga2 during pheromone signaling. Mating factor activates the Fus3/MAPK kinase, whose substrates induce gene expression, cell cycle arrest, and formation of the mating projection. Our studies demonstrate that Fus3 also phosphorylates Rga2 at inhibitory S/TP sites, which are targeted by Cdks during the cell cycle, and that calcineurin opposes Fus3 to activate Rga2 and decrease Cdc42 signaling. Yeast expressing an Rga2 mutant that is defective for regulation by calcineurin display increased gene expression in response to pheromone. This work is the first to identify cross-talk between Ca2+/calcineurin and Cdc42 signaling and to demonstrate modulation of Cdc42 activity through a GAP during mating.

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Phospholipase D1 is required for efficient mating projection formation in

FEMS Yeast Research ◽

10.1016/s1567-1356(01)00027-7 ◽

2001 ◽

Vol 1 (3) ◽

pp. 225-232 ◽

Cited By ~ 1

Author(s):

M HAIRFIELD ◽

A AYERS ◽

J DOLAN

Keyword(s):

Phospholipase D1 ◽

Mating Projection

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The Cbk1p Pathway Is Important for Polarized Cell Growth and Cell Separation in Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.21.7.2449-2462.2001 ◽

2001 ◽

Vol 21 (7) ◽

pp. 2449-2462 ◽

Cited By ~ 128

Author(s):

Scott Bidlingmaier ◽

Eric L. Weiss ◽

Chris Seidel ◽

David G. Drubin ◽

Michael Snyder

Keyword(s):

Cell Wall ◽

Transcriptional Repression ◽

Cell Separation ◽

Apical Growth ◽

Transcriptional Analysis ◽

Gene Encoding ◽

Bud Neck ◽

Encoding Gene ◽

Diploid Cells ◽

Mating Projection

ABSTRACT During the early stages of budding, cell wall remodeling and polarized secretion are concentrated at the bud tip (apical growth). The CBK1 gene, encoding a putative serine/threonine protein kinase, was identified in a screen designed to isolate mutations that affect apical growth. Analysis of cbk1Δ cells reveals that Cbk1p is required for efficient apical growth, proper mating projection morphology, bipolar bud site selection in diploid cells, and cell separation. Epitope-tagged Cbk1p localizes to both sides of the bud neck in late anaphase, just prior to cell separation.CBK1 and another gene, HYM1, were previously identified in a screen for genes involved in transcriptional repression and proposed to function in the same pathway. Deletion ofHYM1 causes phenotypes similar to those observed incbk1Δ cells and disrupts the bud neck localization of Cbk1p. Whole-genome transcriptional analysis of cbk1Δsuggests that the kinase regulates the expression of a number of genes with cell wall-related functions, including two genes required for efficient cell separation: the chitinase-encoding gene CTS1and the glucanase-encoding gene SCW11. The Ace2p transcription factor is required for expression of CTS1 and has been shown to physically interact with Cbk1p. Analysis oface2Δ cells reveals that Ace2p is required for cell separation but not for polarized growth. Our results suggest that Cbk1p and Hym1p function to regulate two distinct cell morphogenesis pathways: an ACE2-independent pathway that is required for efficient apical growth and mating projection formation and anACE2-dependent pathway that is required for efficient cell separation following cytokinesis. Cbk1p is most closely related to theNeurospora crassa Cot-1; Schizosaccharomyces pombe Orb6; Caenorhabditis elegans, Drosophila, and human Ndr; and Drosophila and mammalian WARTS/LATS kinases. Many Cbk1-related kinases have been shown to regulate cellular morphology.

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Pneumocystis carinii Cell Wall Biosynthesis Kinase Gene CBK1 Is an Environmentally Responsive Gene That Complements Cell Wall Defects of cbk-Deficient Yeast

Infection and Immunity ◽

10.1128/iai.72.8.4628-4636.2004 ◽

2004 ◽

Vol 72 (8) ◽

pp. 4628-4636 ◽

Cited By ~ 26

Author(s):

Theodore J. Kottom ◽

Andrew H. Limper

Keyword(s):

Cell Wall ◽

Pneumocystis Carinii ◽

Life Forms ◽

Lung Epithelial Cells ◽

Yeast Cells ◽

Cell Wall Biosynthesis ◽

Kinase Gene ◽

Environmentally Responsive ◽

Responsive Gene ◽

Mating Projection

ABSTRACT Pneumocystis species remain an important cause of life-threatening pneumonia in immunocompromised hosts, including those with AIDS. Responses of the organism to environmental cues both within the lung and elsewhere have been poorly defined. Herein, we report the identification of a cell wall biosynthesis kinase gene (CBK1) homologue in Pneumocystis carinii, isolated by differential display PCR, that is expressed optimally at physiological pH (7 to 8) as opposed to more acidic environments. Expression of Pneumocystis CBK1 was also induced by contact with lung epithelial cells and extracellular matrix. Translation of this gene revealed extensive homology to other fungal CBK1 kinases. Pneumocystis CBK1 expression was equal in the cyst and trophic life forms of the organisms. We further demonstrate that Pneumocystis CBK1 expressed in cbk1Δ Saccharomyces cerevisiae cells restored defective cell wall separation during proliferation. Consistent with this, Pneumocystis CBK1 expression also stimulated transcription of the CTS1 chitinase in cbk1Δ mutant yeast cells, an event necessary for cell wall separation. In addition, Pneumocystis CBK1 cDNA supported normal mating projection formation in response to α-factor in the cbk1Δ yeast cells. Site-directed mutations of serine-303 and threonine-494, potential regulatory phosphorylation sites in Pneumocystis CBK1, abolished mating projection formation, indicating a role for these amino acid residues in CBK1 activity. These findings indicate that Pneumocystis CBK1 is an environmentally responsive gene that may function in signaling pathways necessary for cell growth and mating.

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Polarization of the Yeast Pheromone Receptor Requires Its Internalization but Not Actin-dependent Secretion

Molecular Biology of the Cell ◽

10.1091/mbc.e09-08-0706 ◽

2010 ◽

Vol 21 (10) ◽

pp. 1737-1752 ◽

Cited By ~ 30

Author(s):

Dmitry V. Suchkov ◽

Reagan DeFlorio ◽

Edward Draper ◽

Amber Ismael ◽

Madhushalini Sukumar ◽

...

Keyword(s):

Plasma Membrane ◽

G Protein ◽

Protein Localization ◽

Receptor Internalization ◽

Gradient Sensing ◽

Pheromone Receptor ◽

Surface Receptors ◽

Chemical Gradients ◽

Actin Cables ◽

Mating Projection

In the best understood models of eukaryotic directional sensing, chemotactic cells maintain a uniform distribution of surface receptors even when responding to chemical gradients. The yeast pheromone receptor is also uniformly distributed on the plasma membrane of vegetative cells, but pheromone induces its polarization into “crescents” that cap the future mating projection. Here, we find that in pheromone-treated cells, receptor crescents are visible before detectable polarization of actin cables and that the receptor can polarize in the absence of actin-dependent directed secretion. Receptor internalization, in contrast, seems to be essential for the generation of receptor polarity, and mutations that deregulate this process confer dramatic defects in directional sensing. We also show that pheromone induces the internalization and subsequent polarization of the mating-specific Gα and Gβ proteins and that the changes in G protein localization depend on receptor internalization and receptor–Gα coupling. Our data suggest that the polarization of the receptor and its G protein precedes actin polarization and is important for gradient sensing. We propose that the establishment of receptor/G protein polarity depends on a novel mechanism involving differential internalization and that this serves to amplify the shallow gradient of activated receptor across the cell.

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Polarized Morphogenesis Regulator Spa2 Is Required for the Function of Putative Stretch-Activated Ca2+-Permeable Channel Component Mid1 in Saccharomyces cerevisiae

Eukaryotic Cell ◽

10.1128/ec.4.8.1353-1363.2005 ◽

2005 ◽

Vol 4 (8) ◽

pp. 1353-1363 ◽

Cited By ~ 4

Author(s):

Shigeko Noma ◽

Kazuko Iida ◽

Hidetoshi Iida

Keyword(s):

Double Mutant ◽

Fluorescent Protein ◽

Mitogen Activated Protein Kinase ◽

Peanut Shell ◽

Multicopy Plasmid ◽

Mating Pheromone ◽

Multicopy Suppressor ◽

Amino Terminal ◽

Green Fluorescent ◽

Mating Projection

ABSTRACT Mid1 is a putative stretch-activated Ca2+ channel component and is required for the maintenance of viability in the mating process. In response to mating pheromone, the mid1 mutant normally forms a pointed mating projection but eventually dies. This phenotype is called the mid phenotype. To identify a protein regulating Mid1 or regulated by Mid1, we isolated a multicopy suppressor that rescues the mid1-1 mutant from mating pheromone-induced death and found that it encodes a truncated Spa2 protein lacking an amino-terminal region responsible for interaction with components of the mitogen-activated protein kinase cascades. One of these SPA2 alleles was SPA2ΔN, whose product lacked the region from Ser5 to Leu230. SPA2ΔN on a multicopy plasmid (YEpSPA2ΔN) complemented the mid phenotype but not another phenotype, low Ca2+ accumulation, of the mid1-1 mutant. Neither SPA2ΔN on a low-copy plasmid nor wild-type SPA2 on a multicopy plasmid had suppressive activity. The SPA2 gene is involved in the formation of a pointed mating projection, and cells of the spa2Δ mutant lacking Spa2 are viable and develop a peanut shell-like structure when exposed to mating pheromone. Like the spa2Δ mutant, the mid1-1 spa2Δ double mutant and the mid1-1/YEpSPA2ΔN strain developed the peanut shell-like structure. The mid1-1 spa2Δ double mutant did not have the mid phenotype, indicating that SPA2 is epistatic to MID1. Overexpression of Spa2ΔN abolished the localization of Spa2-green fluorescent protein to the tip of the mating projection. These results suggest that the Spa2ΔN protein interferes with the localization of the normal Spa2 protein and thereby prevents cells from entering the mating process. Therefore, we suggest that Mid1 function is influenced by Spa2 function through polarized morphogenesis.

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Mate and fuse: how yeast cells do it

Open Biology ◽

10.1098/rsob.130008 ◽

2013 ◽

Vol 3 (3) ◽

pp. 130008 ◽

Cited By ~ 126

Author(s):

Laura Merlini ◽

Omaya Dudin ◽

Sophie G. Martin

Keyword(s):

Saccharomyces Cerevisiae ◽

Cellular Response ◽

Current Knowledge ◽

Yeast Cells ◽

Mating Response ◽

Biological Problem ◽

Species Specific ◽

Ascomycete Yeast ◽

Mating Projection ◽

Pheromone Perception

Many cells are able to orient themselves in a non-uniform environment by responding to localized cues. This leads to a polarized cellular response, where the cell can either grow or move towards the cue source. Fungal haploid cells secrete pheromones to signal mating, and respond by growing a mating projection towards a potential mate. Upon contact of the two partner cells, these fuse to form a diploid zygote. In this review, we present our current knowledge on the processes of mating signalling, pheromone-dependent polarized growth and cell fusion in Saccharomyces cerevisiae and Schizosaccharomyces pombe , two highly divergent ascomycete yeast models. While the global architecture of the mating response is very similar between these two species, they differ significantly both in their mating physiologies and in the molecular connections between pheromone perception and downstream responses. The use of both yeast models helps enlighten both conserved solutions and species-specific adaptations to a general biological problem.

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