scholarly journals TRIP13PCH-2 promotes Mad2 localization to unattached kinetochores in the spindle checkpoint response

2015 ◽  
Vol 211 (3) ◽  
pp. 503-516 ◽  
Author(s):  
Christian R. Nelson ◽  
Tom Hwang ◽  
Pin-Hsi Chen ◽  
Needhi Bhalla
Keyword(s):  
Cell Division ◽  
Caenorhabditis Elegans ◽  
Spindle Checkpoint ◽  
Checkpoint Activation ◽  
Checkpoint Signaling ◽  
Checkpoint Response ◽  
Aaa Atpase ◽  
C Elegans ◽  
Anaphase Onset ◽  
Data Support

The spindle checkpoint acts during cell division to prevent aneuploidy, a hallmark of cancer. During checkpoint activation, Mad1 recruits Mad2 to kinetochores to generate a signal that delays anaphase onset. Yet, whether additional factors contribute to Mad2’s kinetochore localization remains unclear. Here, we report that the conserved AAA+ ATPase TRIP13PCH-2 localizes to unattached kinetochores and is required for spindle checkpoint activation in Caenorhabditis elegans. pch-2 mutants effectively localized Mad1 to unattached kinetochores, but Mad2 recruitment was significantly reduced. Furthermore, we show that the C. elegans orthologue of the Mad2 inhibitor p31(comet)CMT-1 interacts with TRIP13PCH-2 and is required for its localization to unattached kinetochores. These factors also genetically interact, as loss of p31(comet)CMT-1 partially suppressed the requirement for TRIP13PCH-2 in Mad2 localization and spindle checkpoint signaling. These data support a model in which the ability of TRIP13PCH-2 to disassemble a p31(comet)/Mad2 complex, which has been well characterized in the context of checkpoint silencing, is also critical for spindle checkpoint activation.

scholarly journals Systematic Analysis in Caenorhabditis elegans Reveals that the Spindle Checkpoint Is Composed of Two Largely Independent Branches

2009 ◽  
Vol 20 (4) ◽  
pp. 1252-1267 ◽  
Author(s):  
Anthony Essex ◽  
Alexander Dammermann ◽  
Lindsay Lewellyn ◽  
Karen Oegema ◽  
Arshad Desai
Keyword(s):  
Caenorhabditis Elegans ◽  
Spindle Checkpoint ◽  
Checkpoint Activation ◽  
Systematic Analysis ◽  
Checkpoint Signaling ◽  
Epistasis Analysis ◽  
Kinetochore Assembly ◽  
Anaphase Onset ◽  
Monopolar Spindle ◽  
Spindle Microtubules

Kinetochores use the spindle checkpoint to delay anaphase onset until all chromosomes have formed bipolar attachments to spindle microtubules. Here, we use controlled monopolar spindle formation to systematically define the requirements for spindle checkpoint signaling in the Caenorhabditis elegans embryo. The results, when interpreted in light of kinetochore assembly epistasis analysis, indicate that checkpoint activation is coordinately directed by the NDC-80 complex, the Rod/Zwilch/Zw10 complex, and BUB-1—three components independently targeted to the outer kinetochore by the scaffold protein KNL-1. These components orchestrate the integration of a core Mad1MDF-1/Mad2MDF-2-based signal, with a largely independent Mad3SAN-1/BUB-3 pathway. Evidence for independence comes from the fact that subtly elevating Mad2MDF-2 levels bypasses the requirement for BUB-3 and Mad3SAN-1 in kinetochore-dependent checkpoint activation. Mad3SAN-1 does not accumulate at unattached kinetochores and BUB-3 kinetochore localization is independent of Mad2MDF-2. We discuss the rationale for a bipartite checkpoint mechanism in which a core Mad1MDF-1/Mad2MDF-2 signal generated at kinetochores is integrated with a separate cytoplasmic Mad3SAN-1/BUB-3–based pathway.

scholarly journals Checkpoint silencing during the DNA damage response in Caenorhabditis elegans embryos

2006 ◽  
Vol 172 (7) ◽  
pp. 999-1008 ◽  
Author(s):  
Antonia H. Holway ◽  
Seung-Hwan Kim ◽  
Adriana La Volpe ◽  
W. Matthew Michael
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Cell Division ◽  
Caenorhabditis Elegans ◽  
Replication Fork ◽  
Germ Line ◽  
Checkpoint Activation ◽  
Checkpoint Response ◽  
Damage Response ◽  
Fork Stalling

In most cells, the DNA damage checkpoint delays cell division when replication is stalled by DNA damage. In early Caenorhabditis elegans embryos, however, the checkpoint responds to developmental signals that control the timing of cell division, and checkpoint activation by nondevelopmental inputs disrupts cell cycle timing and causes embryonic lethality. Given this sensitivity to inappropriate checkpoint activation, we were interested in how embryos respond to DNA damage. We demonstrate that the checkpoint response to DNA damage is actively silenced in embryos but not in the germ line. Silencing requires rad-2, gei-17, and the polh-1 translesion DNA polymerase, which suppress replication fork stalling and thereby eliminate the checkpoint-activating signal. These results explain how checkpoint activation is restricted to developmental signals during embryogenesis and insulated from DNA damage. They also show that checkpoint activation is not an obligatory response to DNA damage and that pathways exist to bypass the checkpoint when survival depends on uninterrupted progression through the cell cycle.

scholarly journals Kinetochore-localized BUB-1/BUB-3 complex promotes anaphase onset in C. elegans

2015 ◽  
Vol 209 (4) ◽  
pp. 507-517 ◽  
Author(s):  
Taekyung Kim ◽  
Mark W. Moyle ◽  
Pablo Lara-Gonzalez ◽  
Christian De Groot ◽  
Karen Oegema ◽  
...  
Keyword(s):  
Spindle Checkpoint ◽  
Kinase Domain ◽  
Mitotic Chromosomes ◽  
Checkpoint Signaling ◽  
C Elegans ◽  
Anaphase Onset ◽  
Checkpoint Pathway ◽  
Chromosome Alignment ◽  
Kinetochore Region

The conserved Bub1/Bub3 complex is recruited to the kinetochore region of mitotic chromosomes, where it initiates spindle checkpoint signaling and promotes chromosome alignment. Here we show that, in contrast to the expectation for a checkpoint pathway component, the BUB-1/BUB-3 complex promotes timely anaphase onset in Caenorhabditis elegans embryos. This activity of BUB-1/BUB-3 was independent of spindle checkpoint signaling but required kinetochore localization. BUB-1/BUB-3 inhibition equivalently delayed separase activation and other events occurring during mitotic exit. The anaphase promotion function required BUB-1’s kinase domain, but not its kinase activity, and this function was independent of the role of BUB-1/BUB-3 in chromosome alignment. These results reveal an unexpected role for the BUB-1/BUB-3 complex in promoting anaphase onset that is distinct from its well-studied functions in checkpoint signaling and chromosome alignment, and suggest a new mechanism contributing to the coordination of the metaphase-to-anaphase transition.

scholarly journals A sequential multi-target Mps1 phosphorylation cascade promotes spindle checkpoint signaling

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Zhejian Ji ◽  
Haishan Gao ◽  
Luying Jia ◽  
Bing Li ◽  
Hongtao Yu
Keyword(s):  
Spindle Checkpoint ◽  
Mitotic Checkpoint ◽  
Anaphase Promoting Complex ◽  
Checkpoint Activation ◽  
Checkpoint Signaling ◽  
Microtubule Attachment ◽  
Anaphase Onset ◽  
Phosphorylation Cascade ◽  
Mitotic Checkpoint Complex

The master spindle checkpoint kinase Mps1 senses kinetochore-microtubule attachment and promotes checkpoint signaling to ensure accurate chromosome segregation. The kinetochore scaffold Knl1, when phosphorylated by Mps1, recruits checkpoint complexes Bub1–Bub3 and BubR1–Bub3 to unattached kinetochores. Active checkpoint signaling ultimately enhances the assembly of the mitotic checkpoint complex (MCC) consisting of BubR1–Bub3, Mad2, and Cdc20, which inhibits the anaphase-promoting complex or cyclosome bound to Cdc20 (APC/CCdc20) to delay anaphase onset. Using in vitro reconstitution, we show that Mps1 promotes APC/C inhibition by MCC components through phosphorylating Bub1 and Mad1. Phosphorylated Bub1 binds to Mad1–Mad2. Phosphorylated Mad1 directly interacts with Cdc20. Mutations of Mps1 phosphorylation sites in Bub1 or Mad1 abrogate the spindle checkpoint in human cells. Therefore, Mps1 promotes checkpoint activation through sequentially phosphorylating Knl1, Bub1, and Mad1. This sequential multi-target phosphorylation cascade makes the checkpoint highly responsive to Mps1 and to kinetochore-microtubule attachment.

scholarly journals The Caenorhabditis elegans Kinetochore Reorganizes at Prometaphase and in Response to Checkpoint Stimuli

2004 ◽  
Vol 15 (11) ◽  
pp. 5187-5196 ◽  
Author(s):  
Jeffrey H. Stear ◽  
Mark B. Roth
Keyword(s):  
Rna Interference ◽  
Caenorhabditis Elegans ◽  
Spindle Checkpoint ◽  
Metaphase Plate ◽  
Gene Products ◽  
Checkpoint Activation ◽  
Checkpoint Response ◽  
Microtubule Attachment

Previous studies of the kinetochore in mammalian systems have demonstrated that this structure undergoes reorganizations after microtubule attachment or in response to activation of the spindle checkpoint. Here, we show that the Caenorhabditis elegans kinetochore displays analogous rearrangements at prometaphase, when microtubule/chromosome interactions are being established, and after exposure to checkpoint stimuli such as nocodazole or anoxia. These reorganizations are characterized by a dissociation of several kinetochore proteins, including HCP-1/CeCENP-F, HIM-10/CeNuf2, SAN-1/CeMad3, and CeBUB-1, from the centromere. We further demonstrate that at metaphase, despite having dissociated from the centromere, these reorganized kinetochore proteins maintain their associations with the metaphase plate. After checkpoint activation, these proteins are detectable as large “flares” that project out laterally from the metaphase plate. Disrupting these gene products via RNA interference results in sensitivity to checkpoint stimuli, as well as defects in the organization of chromosomes at metaphase. These phenotypes suggest that these proteins, and by extension their reorganization during mitosis, are important for mediating the checkpoint response as well as directing the assembly of the metaphase plate.

scholarly journals Regulation of Kinetochore Recruitment of Two Essential Mitotic Spindle Checkpoint Proteins by Mps1 Phosphorylation

2009 ◽  
Vol 20 (1) ◽  
pp. 10-20 ◽  
Author(s):  
Quanbin Xu ◽  
Songcheng Zhu ◽  
Wei Wang ◽  
Xiaojuan Zhang ◽  
William Old ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Mitotic Spindle ◽  
Kinase Activity ◽  
Spindle Checkpoint ◽  
Signaling Cascade ◽  
Checkpoint Activation ◽  
Mitotic Spindle Checkpoint ◽  
Checkpoint Signaling ◽  
Checkpoint Proteins ◽  
Anaphase Onset

Mps1 is a protein kinase that plays essential roles in spindle checkpoint signaling. Unattached kinetochores or lack of tension triggers recruitment of several key spindle checkpoint proteins to the kinetochore, which delays anaphase onset until proper attachment or tension is reestablished. Mps1 acts upstream in the spindle checkpoint signaling cascade, and kinetochore targeting of Mps1 is required for subsequent recruitment of Mad1 and Mad2 to the kinetochore. The mechanisms that govern recruitment of Mps1 or other checkpoint proteins to the kinetochore upon spindle checkpoint activation are incompletely understood. Here, we demonstrate that phosphorylation of Mps1 at T12 and S15 is required for Mps1 recruitment to the kinetochore. Mps1 kinetochore recruitment requires its kinase activity and autophosphorylation at T12 and S15. Mutation of T12 and S15 severely impairs its kinetochore association and markedly reduces recruitment of Mad2 to the kinetochore. Our studies underscore the importance of Mps1 autophosphorylation in kinetochore targeting and spindle checkpoint signaling.

scholarly journals Direct requirement for Xmus101 in ATR-mediated phosphorylation of Claspin bound Chk1 during checkpoint signaling

2006 ◽  
Vol 173 (2) ◽  
pp. 181-186 ◽  
Author(s):  
Shan Yan ◽  
Howard D. Lindsay ◽  
W. Matthew Michael
Keyword(s):  
Sperm Chromatin ◽  
Checkpoint Control ◽  
Direct Role ◽  
Checkpoint Activation ◽  
Replication Checkpoint ◽  
Checkpoint Signaling ◽  
Checkpoint Response ◽  
Egg Extracts ◽  
Oligonucleotide Duplex ◽  
Stalled Replication Forks

TopBP1-like proteins, which include Xenopus laevis Xmus101, are required for DNA replication and have been linked to replication checkpoint control. A direct role for TopBP1/Mus101 in checkpoint control has been difficult to prove, however, because of the requirement for replication in generating the DNA structures that activate the checkpoint. Checkpoint activation occurs in X. laevis egg extracts upon addition of an oligonucleotide duplex (AT70). We show that AT70 bypasses the requirement for replication in checkpoint activation. We take advantage of this replication-independent checkpoint system to determine the role of Xmus101 in the checkpoint. We find that Xmus101 is essential for AT70-mediated checkpoint signaling and that it functions to promote phosphorylation of Claspin bound Chk1 by the ataxia-telangiectasia and Rad-3–related (ATR) protein kinase. We also identify a separation-of-function mutant of Xmus101. In extracts expressing this mutant, replication of sperm chromatin occurs normally; however, the checkpoint response to stalled replication forks fails. These data demonstrate that Xmus101 functions directly during signal relay from ATR to Chk1.

scholarly journals The ESCRT protein Chmp4c regulates mitotic spindle checkpoint signaling

2018 ◽  
Vol 217 (3) ◽  
pp. 861-876 ◽  
Author(s):  
Eleni Petsalaki ◽  
Maria Dandoulaki ◽  
George Zachos
Keyword(s):  
Mitotic Spindle ◽  
Spindle Checkpoint ◽  
Metaphase Plate ◽  
Membrane Remodeling ◽  
Mitotic Progression ◽  
Mitotic Spindle Checkpoint ◽  
Checkpoint Signaling ◽  
Checkpoint Proteins ◽  
Endosomal Sorting ◽  
Anaphase Onset

The mitotic spindle checkpoint delays anaphase onset in the presence of unattached kinetochores, and efficient checkpoint signaling requires kinetochore localization of the Rod–ZW10–Zwilch (RZZ) complex. In the present study, we show that human Chmp4c, a protein involved in membrane remodeling, localizes to kinetochores in prometaphase but is reduced in chromosomes aligned at the metaphase plate. Chmp4c promotes stable kinetochore–microtubule attachments and is required for proper mitotic progression, faithful chromosome alignment, and segregation. Depletion of Chmp4c diminishes localization of RZZ and Mad1-Mad2 checkpoint proteins to prometaphase kinetochores and impairs mitotic arrest when microtubules are depolymerized by nocodazole. Furthermore, Chmp4c binds to ZW10 through a small C-terminal region, and constitutive Chmp4c kinetochore targeting causes a ZW10-dependent checkpoint metaphase arrest. In addition, Chmp4c spindle functions do not require endosomal sorting complex required for transport–dependent membrane remodeling. These results show that Chmp4c regulates the mitotic spindle checkpoint by promoting localization of the RZZ complex to unattached kinetochores.

scholarly journals The Na+-K+-ATPase is needed in glia of touch receptors for responses to touch in C. elegans

2020 ◽  
Vol 123 (5) ◽  
pp. 2064-2074 ◽  
Author(s):  
Christina K. Johnson ◽  
Jesus Fernandez-Abascal ◽  
Ying Wang ◽  
Lei Wang ◽  
Laura Bianchi
Keyword(s):  
Caenorhabditis Elegans ◽  
Avoidance Behavior ◽  
Molecular Mechanisms ◽  
C Elegans ◽  
Data Support ◽  
Accessory Cells ◽  
Touch Avoidance ◽  
Neuronal Output

Increasing evidences support that accessory cells in mechanosensors regulate neuronal output; however, the glial molecular mechanisms that control this regulation are not fully understood. We show here in Caenorhabditis elegans that specific glial Na+-K+-ATPase genes are needed for nose touch-avoidance behavior. Our data support the requirement of these Na+-K+-ATPases for homeostasis of Na+ and K+ in nose touch receptors. Our data add to our understanding of glial regulation of mechanosensors.

scholarly journals Plx1 is the 3F3/2 kinase responsible for targeting spindle checkpoint proteins to kinetochores

2005 ◽  
Vol 170 (5) ◽  
pp. 709-719 ◽  
Author(s):  
Oi Kwan Wong ◽  
Guowei Fang
Keyword(s):  
Spindle Checkpoint ◽  
Cellular Responses ◽  
High Fidelity ◽  
Checkpoint Response ◽  
Checkpoint Proteins ◽  
Tension Response ◽  
Anaphase Onset ◽  
Chromosome Separation ◽  
Pulling Force ◽  
Necessary And Sufficient

Dynamic attachment of microtubules to kinetochores during mitosis generates pulling force, or tension, required for the high fidelity of chromosome separation. A lack of tension activates the spindle checkpoint and delays the anaphase onset. A key step in the tension–response pathway involves the phosphorylation of the 3F3/2 epitope by an unknown kinase on untensed kinetochores. Using a rephosphorylation assay in Xenopus laevis extracts, we identified the kinetochore-associated Polo-like kinase Plx1 as the kinase both necessary and sufficient for this phosphorylation. Indeed, Plx1 is the physiological 3F3/2 kinase involved in checkpoint response, as immunodepletion of Plx1 from checkpoint extracts abolished the 3F3/2 signal and blocked association of xMad2, xBubR1, xNdc80, and xNuf2 with kinetochores. Interestingly, the kinetochore localization of Plx1 is under the control of the checkpoint protein xMps1, as immunodepletion of xMps1 prevents binding of Plx1 to kinetochores. Thus, Plx1 couples the tension signal to cellular responses through phosphorylating the 3F3/2 epitope and targeting structural and checkpoint proteins to kinetochores.

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