scholarly journals Targeting and functional mechanisms of the cytokinesis‑related F‑BAR protein Hof1 during the cell cycle

2013 ◽  
Vol 24 (9) ◽  
pp. 1305-1320 ◽  
Author(s):  
Younghoon Oh ◽  
Jennifer Schreiter ◽  
Ryuichi Nishihama ◽  
Carsten Wloka ◽  
Erfei Bi
Keyword(s):  
Cell Cycle ◽  
Budding Yeast ◽  
Coiled Coil ◽  
Dependent Manner ◽  
Polarized Growth ◽  
Yeast Saccharomyces Cerevisiae ◽  
Septum Formation ◽  
Primary Septum ◽  
Bud Neck ◽  
Ring Constriction

F-BAR proteins are membrane‑associated proteins believed to link the plasma membrane to the actin cytoskeleton in cellular processes such as cytokinesis and endocytosis. In the budding yeast Saccharomyces cerevisiae, the F‑BAR protein Hof1 localizes to the division site in a complex pattern during the cell cycle and plays an important role in cytokinesis. However, the mechanisms underlying its localization and function are poorly understood. Here we show that Hof1 contains three distinct targeting domains that contribute to cytokinesis differentially. The N‑terminal half of Hof1 localizes to the bud neck and the sites of polarized growth during the cell cycle. The neck localization is mediated mainly by an interaction between the second coiled‑coil region in the N‑terminus and the septin Cdc10, whereas the localization to the sites of polarized growth is mediated entirely by the F‑BAR domain. In contrast, the C‑terminal half of Hof1 interacts with Myo1, the sole myosin‑II heavy chain in budding yeast, and localizes to the bud neck in a Myo1‑dependent manner from the onset to the completion of cytokinesis. We also show that the SH3 domain in the C‑terminus plays an important role in maintaining the symmetry of Myo1 ring constriction during cytokinesis and that Hof1 interacts with Chs2, a chitin synthase that is required for primary septum formation. Together these data define a mechanism that accounts for the localization of Hof1 during the cell cycle and suggest that Hof1 may function in cytokinesis by coupling actomyosin ring constriction to primary septum formation through interactions with Myo1 and Chs2.

scholarly journals The BUD4 protein of yeast, required for axial budding, is localized to the mother/BUD neck in a cell cycle-dependent manner.

1996 ◽  
Vol 134 (2) ◽  
pp. 413-427 ◽  
Author(s):  
S L Sanders ◽  
I Herskowitz
Keyword(s):  
Cell Cycle ◽  
Binding Motif ◽  
Temperature Sensitive ◽  
Dependent Manner ◽  
Alpha Cells ◽  
Yeast Saccharomyces Cerevisiae ◽  
Bud Neck ◽  
A Cell ◽  
Cycle Dependent ◽  
Diploid Cells

A and alpha cells of the yeast Saccharomyces cerevisiae exhibit an axial budding pattern, whereas a/alpha diploid cells exhibit a bipolar pattern. Mutations in BUD3, BUD4, and AXL1 cause a and alpha cells to exhibit the bipolar pattern, indicating that these genes are necessary to specify the axial budding pattern (Chant, J., and I. Herskowitz. 1991. Cell. 65:1203-1212; Fujita, A., C. Oka, Y. Arikawa, T. Katagi, A. Tonouchi, S. Kuhara, and Y. Misumi. 1994. Nature (Lond.). 372:567-570). We cloned and sequenced BUD4, which codes for a large, novel protein (Bud4p) with a potential GTP-binding motif. Bud4p is expressed and localized to the mother/bud neck in all cell types. Most mitotic cells contain two apparent rings of Bud4 immunoreactive staining, as observed for Bud3p (Chant, J., M. Mischke, E. Mitchell, I. Herskowitz, and J.R. Pringle. 1995. J. Cell Biol. 129: 767-778). Early G1 cells contain a single ring of Bud4p immunoreactive staining, whereas cells at START and in S phase lack these rings. The level of Bud4p is also regulated in a cell cycle-dependent manner. Bud4p is inefficiently localized in bud3 mutants and after a temperature shift of a temperature-sensitive mutant, cdc12, defective in the neck filaments. These observations suggest that Bud4p and Bud3p cooperate to recognize a spatial landmark (the neck filaments) during mitosis and support the hypothesis that they subsequently become a landmark for establishing the axial budding pattern in G1.

scholarly journals Mitotic exit kinase Dbf2 directly phosphorylates chitin synthase Chs2 to regulate cytokinesis in budding yeast

2012 ◽  
Vol 23 (13) ◽  
pp. 2445-2456 ◽  
Author(s):  
Younghoon Oh ◽  
Kuang-Jung Chang ◽  
Peter Orlean ◽  
Carsten Wloka ◽  
Raymond Deshaies ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Budding Yeast ◽  
Cell Cycle Control ◽  
Mitotic Exit ◽  
Chitin Synthase ◽  
Chitin Synthesis ◽  
Yeast Saccharomyces Cerevisiae ◽  
Ecm Remodeling ◽  
Division Site ◽  
Primary Septum

How cell cycle machinery regulates extracellular matrix (ECM) remodeling during cytokinesis remains poorly understood. In the budding yeast Saccharomyces cerevisiae, the primary septum (PS), a functional equivalent of animal ECM, is synthesized during cytokinesis by the chitin synthase Chs2. Here, we report that Dbf2, a conserved mitotic exit kinase, localizes to the division site after Chs2 and directly phosphorylates Chs2 on several residues, including Ser-217. Both phosphodeficient (chs2‑S217A) and phosphomimic (chs2‑S217D) mutations cause defects in cytokinesis, suggesting that dynamic phosphorylation–dephosphorylation of Ser-217 is critical for Chs2 function. It is striking that Chs2‑S217A constricts asymmetrically with the actomyosin ring (AMR), whereas Chs2-S217D displays little or no constriction and remains highly mobile at the division site. These data suggest that Chs2 phosphorylation by Dbf2 triggers its dissociation from the AMR during the late stage of cytokinesis. Of interest, both chs2‑S217A and chs2‑S217D mutants are robustly suppressed by increased dosage of Cyk3, a cytokinesis protein that displays Dbf2‑dependent localization and also stimulates Chs2‑mediated chitin synthesis. Thus Dbf2 regulates PS formation through at least two independent pathways: direct phosphorylation and Cyk3‑mediated activation of Chs2. Our study establishes a mechanism for direct cell cycle control of ECM remodeling during cytokinesis.

scholarly journals Systems Level Modeling of the Cell Cycle Using Budding Yeast

2007 ◽  
Vol 3 ◽  
pp. 117693510700300 ◽  
Author(s):  
B.P. Ingalls ◽  
B.P. Duncker ◽  
D.R. Kim ◽  
B.J. McConkey
Keyword(s):  
Cell Cycle ◽  
Budding Yeast ◽  
Human Cancer ◽  
Quality Data ◽  
Cell Cycle Gene ◽  
Mathematical Framework ◽  
Gene Transcript ◽  
Proteomics Data ◽  
Yeast Saccharomyces Cerevisiae ◽  
Protein Levels

Proteins involved in the regulation of the cell cycle are highly conserved across all eukaryotes, and so a relatively simple eukaryote such as yeast can provide insight into a variety of cell cycle perturbations including those that occur in human cancer. To date, the budding yeast Saccharomyces cerevisiae has provided the largest amount of experimental and modeling data on the progression of the cell cycle, making it a logical choice for in-depth studies of this process. Moreover, the advent of methods for collection of high-throughput genome, transcriptome, and proteome data has provided a means to collect and precisely quantify simultaneous cell cycle gene transcript and protein levels, permitting modeling of the cell cycle on the systems level. With the appropriate mathematical framework and sufficient and accurate data on cell cycle components, it should be possible to create a model of the cell cycle that not only effectively describes its operation, but can also predict responses to perturbations such as variation in protein levels and responses to external stimuli including targeted inhibition by drugs. In this review, we summarize existing data on the yeast cell cycle, proteomics technologies for quantifying cell cycle proteins, and the mathematical frameworks that can integrate this data into representative and effective models. Systems level modeling of the cell cycle will require the integration of high-quality data with the appropriate mathematical framework, which can currently be attained through the combination of dynamic modeling based on proteomics data and using yeast as a model organism.

scholarly journals Far3 and Five Interacting Proteins Prevent Premature Recovery from Pheromone Arrest in the Budding Yeast Saccharomyces cerevisiae

2003 ◽  
Vol 23 (5) ◽  
pp. 1750-1763 ◽  
Author(s):  
Hilary A. Kemp ◽  
George F. Sprague,
Keyword(s):  
Cell Cycle ◽  
Budding Yeast ◽  
Open Reading Frames ◽  
Velocity Sedimentation ◽  
Interacting Proteins ◽  
Yeast Saccharomyces Cerevisiae ◽  
Cycle Arrest ◽  
Mutant Cells ◽  
Two Hybrid ◽  
Reading Frames

ABSTRACT In budding yeast, diffusible mating pheromones initiate a signaling pathway that culminates in several responses, including cell cycle arrest. Only a handful of genes required for the interface between pheromone response and the cell cycle have been identified, among them FAR1 and FAR3; of these, only FAR1 has been extensively characterized. In an effort to learn about the mechanism by which Far3 acts, we used the two-hybrid method to identify interacting proteins. We identified five previously uncharacterized open reading frames, dubbed FAR7, FAR8, FAR9, FAR10, and FAR11, that cause a far3-like pheromone arrest defect when disrupted. Using two-hybrid and coimmunoprecipitation analysis, we found that all six Far proteins interact with each other. Moreover, velocity sedimentation experiments suggest that Far3 and Far7 to Far11 form a complex. The phenotype of a sextuple far3far7-far11 mutant is no more severe than any single mutant. Thus, FAR3 and FAR7 to FAR11 all participate in the same pathway leading to G1 arrest. These mutants initially arrest in response to pheromone but resume budding after 10 h. Under these conditions, wild-type cells fail to resume budding even after several days whereas far1 mutant cells resume budding within 1 h. We conclude that the FAR3-dependent arrest pathway is functionally distinct from that which employs FAR1.

scholarly journals Dynamic Localization of Protein Phosphatase Type 1 in the Mitotic Cell Cycle of Saccharomyces cerevisiae

2000 ◽  
Vol 149 (1) ◽  
pp. 125-140 ◽  
Author(s):  
Andrew Bloecher ◽  
Kelly Tatchell
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Protein Phosphatase ◽  
Essential Gene ◽  
Mitotic Cell ◽  
Type I ◽  
Mitotic Cell Cycle ◽  
Dependent Manner ◽  
Bud Neck ◽  
Protein Phosphatase Type

Protein phosphatase type I (PP1), encoded by the single essential gene GLC7 in Saccharomyces cerevisiae, functions in diverse cellular processes. To identify in vivo subcellular location(s) where these processes take place, we used a functional green fluorescent protein (GFP)–Glc7p fusion protein. Time-lapse fluorescence microscopy revealed GFP–Glc7p localizes predominantly in the nucleus throughout the mitotic cell cycle, with the highest concentrations in the nucleolus. GFP–Glc7p was also observed in a ring at the bud neck, which was dependent upon functional septins. Supporting a role for Glc7p in bud site selection, a glc7-129 mutant displayed a random budding pattern. In α-factor treated cells, GFP–Glc7p was located at the base of mating projections, again in a septin-dependent manner. At the start of anaphase, GFP–Glc7p accumulated at the spindle pole bodies and remained there until cytokinesis. After anaphase, GFP–Glc7p became concentrated in a ring that colocalized with the actomyosin ring. A GFP–Glc7-129 fusion was defective in localizing to the bud neck and SPBs. Together, these results identify sites of Glc7p function and suggest Glc7p activity is regulated through dynamic changes in its location.

Fission yeast Pds5 is required for accurate chromosome segregation and for survival after DNA damage or metaphase arrest

2002 ◽  
Vol 115 (3) ◽  
pp. 587-598 ◽  
Author(s):  
Shao-Win Wang ◽  
Rebecca L. Read ◽  
Chris J. Norbury
Keyword(s):  
Dna Damage ◽  
Fission Yeast ◽  
Sister Chromatid ◽  
Budding Yeast ◽  
Eukaryotic Cell ◽  
Sister Chromatid Cohesion ◽  
Chromosome Loss ◽  
Dependent Manner ◽  
Yeast Saccharomyces Cerevisiae ◽  
Chromatid Cohesion

Sister chromatid cohesion, which is established during the S phase of the eukaryotic cell cycle and persists until the onset of anaphase, is essential for the maintenance of genomic integrity. Cohesion requires the multi-protein complex cohesin, as well as a number of accessory proteins including Pds5/BIMD/Spo76. In the budding yeast Saccharomyces cerevisiae Pds5 is an essential protein that localises to chromosomes in a cohesin-dependent manner. Here we describe the characterisation in the fission yeast Schizosaccharomyces pombe of pds5+, a novel,non-essential orthologue of S. cerevisiae PDS5. The S. pombePds5 protein was localised to punctate nuclear foci in a manner that was dependent on the Rad21 cohesin component. This, together with additional genetic evidence, points towards an involvement of S. pombe Pds5 in sister chromatid cohesion. S. pombe pds5 mutants were hypersensitive to DNA damage and to mitotic metaphase delay, but this sensitivity was apparently not due to precocious loss of sister chromatid cohesion. These cells also suffered increased spontaneous chromosome loss and meiotic defects and their viability was dependent on the spindle checkpoint protein Bub1. Thus, while S. pombe Pds5 has an important cohesin-related role, this differs significantly from that of the equivalent budding yeast protein.

scholarly journals Scw1p Antagonizes the Septation Initiation Network To Regulate Septum Formation and Cell Separation in the Fission Yeast Schizosaccharomyces pombe

2003 ◽  
Vol 2 (3) ◽  
pp. 510-520 ◽  
Author(s):  
Quan-Wen Jin ◽  
Dannel McCollum
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Cell Separation ◽  
Deletion Mutant ◽  
Temperature Sensitive ◽  
Growth Defects ◽  
Septum Formation ◽  
Primary Septum ◽  
Septation Initiation Network ◽  
Ring Constriction

ABSTRACT Cytokinesis in the fission yeast Schizosaccharomyces pombe is regulated by a signaling pathway termed the septation initiation network (SIN). The SIN is essential for initiation of actomyosin ring constriction and septum formation. In a screen to search for mutations that can rescue the sid2-250 SIN mutant, we obtained scw1-18. Both the scw1-18 mutant and the scw1 deletion mutant (scw1Δ mutant), have defects in cell separation. Both the scw1-18 and scw1Δ mutations rescue the growth defects of not just the sid2-250 mutant but also the other temperature-sensitive SIN mutants. Other cytokinesis mutants, such as those defective for actomyosin ring formation, are not rescued by scw1Δ. scw1Δ does not seem to rescue the SIN by restoring SIN signaling defects. However, scw1Δ may function downstream of the SIN to promote septum formation, since scw1Δ can rescue the septum formation defects of the cps1-191β-1,3-glucan synthase mutant, which is required for synthesis of the primary septum.

scholarly journals LIN-5 Is a Novel Component of the Spindle Apparatus Required for Chromosome Segregation and Cleavage Plane Specification in Caenorhabditis elegans

2000 ◽  
Vol 148 (1) ◽  
pp. 73-86 ◽  
Author(s):  
Monique A. Lorson ◽  
H. Robert Horvitz ◽  
Sander van den Heuvel
Keyword(s):  
Cell Cycle ◽  
Caenorhabditis Elegans ◽  
Dna Replication ◽  
Chromosome Segregation ◽  
Cleavage Plane ◽  
Coiled Coil ◽  
Metaphase Plate ◽  
Dependent Manner ◽  
Spindle Apparatus ◽  
Cytoplasmic Cleavage

Successful divisions of eukaryotic cells require accurate and coordinated cycles of DNA replication, spindle formation, chromosome segregation, and cytoplasmic cleavage. The Caenorhabditis elegans gene lin-5 is essential for multiple aspects of cell division. Cells in lin-5 null mutants enter mitosis at the normal time and form bipolar spindles, but fail chromosome alignment at the metaphase plate, sister chromatid separation, and cytokinesis. Despite these defects, cells exit from mitosis without delay and progress through subsequent rounds of DNA replication, centrosome duplication, and abortive mitoses. In addition, early embryos that lack lin-5 function show defects in spindle positioning and cleavage plane specification. The lin-5 gene encodes a novel protein with a central coiled-coil domain. This protein localizes to the spindle apparatus in a cell cycle- and microtubule-dependent manner. The LIN-5 protein is located at the centrosomes throughout mitosis, at the kinetochore microtubules in metaphase cells, and at the spindle during meiosis. Our results show that LIN-5 is a novel component of the spindle apparatus required for chromosome and spindle movements, cytoplasmic cleavage, and correct alternation of the S and M phases of the cell cycle.

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