Title: A Noncanonical Hippo Pathway Regulates Spindle Disassembly and Cytokinesis During Meiosis in Saccharomyces cerevisiae
Abstract Meiosis in the budding yeast Saccharomyces cerevisiae is used to create haploid yeast spores from a diploid mother cell. During meiosis II, cytokinesis occurs by closure of the prospore membrane, a membrane that initiates at the spindle pole body and grows to surround each of the haploid meiotic products. Timely prospore membrane closure requires SPS1, which encodes an STE20 family GCKIII kinase. To identify genes that may activate SPS1, we utilized a histone phosphorylation defect of sps1 mutants to screen for genes with a similar phenotype and found that cdc15 shared this phenotype. CDC15 encodes a Hippo-like kinase that is part of the mitotic exit network. We find that Sps1 complexes with Cdc15, that Sps1 phosphorylation requires Cdc15, and that CDC15 is also required for timely prospore membrane closure. We also find that SPS1, like CDC15, is required for meiosis II spindle disassembly and sustained anaphase II release of Cdc14 in meiosis. However, the NDR-kinase complex encoded by DBF2/DBF20MOB1 which functions downstream of CDC15 in mitotic cells, does not appear to play a role in spindle disassembly, timely prospore membrane closure, or sustained anaphase II Cdc14 release. Taken together, our results suggest that the mitotic exit network is rewired for exit from meiosis II, such that SPS1 replaces the NDR-kinase complex downstream of CDC15. more »« less
Seitz, Brian C.; Mucelli, Xheni; Majano, Maira; Wallis, Zoey; Dodge, Ashley C.; Carmona, Catherine; Durant, Matthew; Maynard, Sharra; Huang, Linda S.
(, Molecular Biology of the Cell)
Bloom, Kerry
(Ed.)
AMA1 and SPS1 control distinct aspects of meiosis II spindle disassembly, with AMA1 affecting the loss of Ase1 and Cin8 during meiosis II spindle disassembly, while SPS1 affects Bim1. The Anaphase Promoting Complex and meiotic/mitotic exit pathways seem to regulate similar targets in meiosis as mitosis, despite utilizing meiosis-specific regulators in those pathways.
Durant, Matthew; Roesner, Joseph M.; Mucelli, Xheni; Slubowski, Christian J.; Klee, Erin; Seitz, Brian C.; Wallis, Zoey; Huang, Linda S.
(, Journal of Fungi)
During sporulation in the budding yeast Saccharomyces cerevisiae, proper development of the prospore membrane is necessary for the formation of viable spores. The prospore membrane will eventually become the plasma membrane of the newly formed haploid spore and also serves as the template for the deposition of the spore wall. The prospore membrane is generated de novo during meiosis II and the growing edge of the prospore membrane is associated with the Leading Edge Protein (LEP) complex. We find that the Smk1 MAP kinase, along with its activator Ssp2, transiently localizes with the LEP during late meiosis II. SSP2 is required for the leading edge localization of Smk1; this localization is independent of the activation state of Smk1. Like other LEP components, the localization of Smk1 at the leading edge also depends on Ady3. Although prospore membrane development begins normally in smk1 and ssp2 mutants, late prospore membrane formation is disrupted, with the formation of ectopic membrane compartments. Thus, MAP kinase signaling plays an important role in the formation of the prospore membrane.
Weiss, Jodi D.; McVey, Shelby L.; Stinebaugh, Sarah E.; Sullivan, Caroline F.; Dawe, R. Kelly; Nannas, Natalie J.
(, International Journal of Molecular Sciences)
The success of an organism is contingent upon its ability to faithfully pass on its genetic material. In the meiosis of many species, the process of chromosome segregation requires that bipolar spindles be formed without the aid of dedicated microtubule organizing centers, such as centrosomes. Here, we describe detailed analyses of acentrosomal spindle assembly and disassembly in time-lapse images, from live meiotic cells of Zea mays. Microtubules organized on the nuclear envelope with a perinuclear ring structure until nuclear envelope breakdown, at which point microtubules began bundling into a bipolar form. However, the process and timing of spindle assembly was highly variable, with frequent assembly errors in both meiosis I and II. Approximately 61% of cells formed incorrect spindle morphologies, with the most prevalent being tripolar spindles. The erroneous spindles were actively rearranged to bipolar through a coalescence of poles before proceeding to anaphase. Spindle disassembly occurred as a two-state process with a slow depolymerization, followed by a quick collapse. The results demonstrate that maize meiosis I and II spindle assembly is remarkably fluid in the early assembly stages, but otherwise proceeds through a predictable series of events.
Jones, Michele H; Gergely, Zachary R; Steckhahn, Daniel; Zhou, Bojun; Betterton, Meredith D
(, Current Biology)
Kinesin-5 motors play an essential role during mitotic spindle assembly in many organisms: they crosslink antiparallel spindle microtubules, step toward plus ends, and slide the microtubules apart. This activity separates the spindle poles and chromosomes. Kinesin-5s are not only plus-end-directed, but can walk or be carried toward MT minus ends where they show enhanced localization. The kinesin-5 C-terminal tail interacts with and regulates the motor, affecting structure, motility, and sliding force of purified kinesin-535–37 along with motility and spindle assembly in cells. The tail contains phosphorylation sites, particularly in the conserved BimC box. Nine mitotic phosphorylation sites were identified in the kinesin-5 motor of the fission yeast Schizosaccharomyces pombe, suggesting that multi-site phosphorylation may regulate kinesin-5s. Here, we show that mutating all nine sites to either alanine or glutamate causes temperature-sensitive lethality due to a failure of bipolar spindle assembly. We characterize kinesin-5 localization and sliding force in the spindle, based on Cut7-dependent microtubule minus-end protrusions in cells lacking kinesin-14 motors. Imaging and computational modeling show that Cut7p simultaneously moves toward minus ends of protrusion MTs and plus ends of spindle midzone MTs. Phosphorylation mutants show dramatic decreases in protrusions and sliding force. Comparison to a model of force to create protrusions suggests that tail truncation and phosphorylation mutants decrease Cut7p sliding force similarly to tail-truncated human Eg5. Our results show that C-terminal tail phosphorylation is required for kinesin-5/Cut7 sliding force and bipolar spindle assembly in fission yeast.
Edelmaier, Christopher; Lamson, Adam R; Gergely, Zachary R; Ansari, Saad; Blackwell, Robert; McIntosh, J Richard; Glaser, Matthew A; Betterton, Meredith D
(, eLife)
The essential functions required for mitotic spindle assembly and chromosome biorientation and segregation are not fully understood, despite extensive study. To illuminate the combinations of ingredients most important to align and segregate chromosomes and simultaneously assemble a bipolar spindle, we developed a computational model of fission-yeast mitosis. Robust chromosome biorientation requires progressive restriction of attachment geometry, destabilization of misaligned attachments, and attachment force dependence. Large spindle length fluctuations can occur when the kinetochore-microtubule attachment lifetime is long. The primary spindle force generators are kinesin-5 motors and crosslinkers in early mitosis, while interkinetochore stretch becomes important after biorientation. The same mechanisms that contribute to persistent biorientation lead to segregation of chromosomes to the poles after anaphase onset. This model therefore provides a framework to interrogate key requirements for robust chromosome biorientation, spindle length regulation, and force generation in the spindle.
Paulissen, Scott M, Hunt, Cindy A, Seitz, Brian C, Slubowski, Christian J, Yu, Yao, Mucelli, Xheni, Truong, Dang, Wallis, Zoey, Nguyen, Hung T, Newman-Toledo, Shayla, Neiman, Aaron M, and Huang, Linda S. A Noncanonical Hippo Pathway Regulates Spindle Disassembly and Cytokinesis During Meiosis in Saccharomyces cerevisiae. Retrieved from https://par.nsf.gov/biblio/10497114. Genetics 216.2 Web. doi:10.1534/genetics.120.303584.
Paulissen, Scott M, Hunt, Cindy A, Seitz, Brian C, Slubowski, Christian J, Yu, Yao, Mucelli, Xheni, Truong, Dang, Wallis, Zoey, Nguyen, Hung T, Newman-Toledo, Shayla, Neiman, Aaron M, & Huang, Linda S. A Noncanonical Hippo Pathway Regulates Spindle Disassembly and Cytokinesis During Meiosis in Saccharomyces cerevisiae. Genetics, 216 (2). Retrieved from https://par.nsf.gov/biblio/10497114. https://doi.org/10.1534/genetics.120.303584
Paulissen, Scott M, Hunt, Cindy A, Seitz, Brian C, Slubowski, Christian J, Yu, Yao, Mucelli, Xheni, Truong, Dang, Wallis, Zoey, Nguyen, Hung T, Newman-Toledo, Shayla, Neiman, Aaron M, and Huang, Linda S.
"A Noncanonical Hippo Pathway Regulates Spindle Disassembly and Cytokinesis During Meiosis in Saccharomyces cerevisiae". Genetics 216 (2). Country unknown/Code not available: Genetics Society of America. https://doi.org/10.1534/genetics.120.303584.https://par.nsf.gov/biblio/10497114.
@article{osti_10497114,
place = {Country unknown/Code not available},
title = {A Noncanonical Hippo Pathway Regulates Spindle Disassembly and Cytokinesis During Meiosis in Saccharomyces cerevisiae},
url = {https://par.nsf.gov/biblio/10497114},
DOI = {10.1534/genetics.120.303584},
abstractNote = {Abstract Meiosis in the budding yeast Saccharomyces cerevisiae is used to create haploid yeast spores from a diploid mother cell. During meiosis II, cytokinesis occurs by closure of the prospore membrane, a membrane that initiates at the spindle pole body and grows to surround each of the haploid meiotic products. Timely prospore membrane closure requires SPS1, which encodes an STE20 family GCKIII kinase. To identify genes that may activate SPS1, we utilized a histone phosphorylation defect of sps1 mutants to screen for genes with a similar phenotype and found that cdc15 shared this phenotype. CDC15 encodes a Hippo-like kinase that is part of the mitotic exit network. We find that Sps1 complexes with Cdc15, that Sps1 phosphorylation requires Cdc15, and that CDC15 is also required for timely prospore membrane closure. We also find that SPS1, like CDC15, is required for meiosis II spindle disassembly and sustained anaphase II release of Cdc14 in meiosis. However, the NDR-kinase complex encoded by DBF2/DBF20MOB1 which functions downstream of CDC15 in mitotic cells, does not appear to play a role in spindle disassembly, timely prospore membrane closure, or sustained anaphase II Cdc14 release. Taken together, our results suggest that the mitotic exit network is rewired for exit from meiosis II, such that SPS1 replaces the NDR-kinase complex downstream of CDC15.},
journal = {Genetics},
volume = {216},
number = {2},
publisher = {Genetics Society of America},
author = {Paulissen, Scott M and Hunt, Cindy A and Seitz, Brian C and Slubowski, Christian J and Yu, Yao and Mucelli, Xheni and Truong, Dang and Wallis, Zoey and Nguyen, Hung T and Newman-Toledo, Shayla and Neiman, Aaron M and Huang, Linda S},
}
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