RZZ and Mad1 dynamics in Drosophila mitosis

Défachelles, Lénaïg; Raich, Natacha; Terracol, Régine; Baudin, Xavier; Williams, Byron; Goldberg, Michael L.; Karess, Roger E.

doi:10.1007/s10577-015-9472-x

Cited by 19 publications

(25 citation statements)

References 45 publications

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“…To determine how Polo activity regulates Rod, we imaged neuroblasts expressing Rod-GFP under the regulation of its endogenous promoter in the presence of either Polo WT or Polo T182D . In accordance with previous studies (Basto et al, 2004;Siller et al, 2005;Défachelles et al, 2015), Rod-GFP localized prominently to KTs during prometaphase and was significantly reduced during metaphase in Polo WT neuroblasts ( Fig 4A-C). Detailed inspection shows that Rod-GFP signal starts to decrease as KTs congress and fades when these stably align at the cell equator ( Fig 4A and B; Movie EV6).…”

Section: Constitutively Active Polo Impairs Rzz Stripping From Late Csupporting

confidence: 92%

Polo regulates Spindly to prevent premature stabilization of kinetochore–microtubule attachments

et al. 2019

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Accurate chromosome segregation in mitosis requires sister kinetochores to bind to microtubules from opposite spindle poles. The stability of kinetochore-microtubule attachments is fine-tuned to prevent or correct erroneous attachments while preserving amphitelic interactions. Polo kinase has been implicated in both stabilizing and destabilizing kinetochore-microtubule attachments. However, the mechanism underlying Polo-destabilizing activity remains elusive. Here, resorting to an RNAi screen in Drosophila for suppressors of a constitutively active Polo mutant, we identified a strong genetic interaction between Polo and the Rod-ZW10-Zwilch (RZZ) complex, whose kinetochore accumulation has been shown to antagonize microtubule stability. We find that Polo phosphorylates Spindly and impairs its ability to bind to Zwilch. This precludes dynein-mediated removal of the RZZ from kinetochores and consequently delays the formation of stable end-on attachments. We propose that high Polo-kinase activity following mitotic entry directs the RZZ complex to minimize premature stabilization of erroneous attachments, whereas a decrease in active Polo in later mitotic stages allows the formation of stable amphitelic spindle attachments. Our findings demonstrate that Polo tightly regulates the RZZ-Spindly-dynein module during mitosis to ensure the fidelity of chromosome segregation.

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Section: Constitutively Active Polo Impairs Rzz Stripping From Late Csupporting

confidence: 92%

Polo regulates Spindly to prevent premature stabilization of kinetochore–microtubule attachments

et al. 2019

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“…Mad1 kinetochore localization is also dependent on the RZZ complex [ 54 – 56 ], and interactions between these components in Drosophila embryos has recently been shown through co-immunoprecipitation experiments [ 57 ]. This complex in turn requires KNL-1 for kinetochore targeting [ 56 , 58 ], although loss of KNL-1 (or Bub1) in mitotically arrested human cells does not completely abolish Mad1 or RZZ kinetochore localization and implicates an additional mechanism for complete recruitment [ 59 ].…”

Section: Discussionmentioning

confidence: 99%

Auxin/AID versus conventional knockouts: distinguishing the roles of CENP-T/W in mitotic kinetochore assembly and stability

et al. 2016

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Most studies using knockout technologies to examine protein function have relied either on shutting off transcription (conventional conditional knockouts with tetracycline-regulated gene expression or gene disruption) or destroying the mature mRNA (RNAi technology). In both cases, the target protein is lost at a rate determined by its intrinsic half-life. Thus, protein levels typically fall over at least 1–3 days, and cells continue to cycle while exposed to a decreasing concentration of the protein. Here we characterise the kinetochore proteome of mitotic chromosomes isolated from a cell line in which the essential kinetochore protein CENP-T is present as an auxin-inducible degron (AID) fusion protein that is fully functional and able to support the viability of the cells. Stripping of the protein from chromosomes in early mitosis via targeted proteasomal degradation reveals the dependency of other proteins on CENP-T for their maintenance in kinetochores. We compare these results with the kinetochore proteome of conventional CENP-T/W knockouts. As the cell cycle is mostly formed from G1, S and G2 phases a gradual loss of CENP-T/W levels is more likely to reflect dependencies associated with kinetochore assembly pre-mitosis and upon entry into mitosis. Interestingly, a putative super-complex involving Rod-Zw10-zwilch (RZZ complex), Spindly, Mad1/Mad2 and CENP-E requires the function of CENP-T/W during kinetochore assembly for its stable association with the outer kinetochore, but once assembled remains associated with chromosomes after stripping of CENP-T during mitosis. This study highlights the different roles core kinetochore components may play in the assembly of kinetochores (upon entry into mitosis) versus the maintenance of specific components (during mitosis).

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“…A candidate is the RZZ complex. RZZ shows classic expansion/compaction behavior, is required for recruitment of several outer-kinetochore components, and has a molecular architecture resembling that of membrane coating proteins that can form polymeric states 25,[35][36][37][38][39][40][41][42][43][44] . We here set out to identify the nature of the fibrous corona, the molecular mechanisms of its ability to expand and compact, and its functional relevance for chromosome segregation.…”

Section: Introductionmentioning

confidence: 99%

Dynamic Kinetochore Size Regulation Promotes Microtubule Capture and Chromosome Biorientation in Mitosis

Sacristán

Ahmad

Fermie

et al. 2018

Preprint

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Faithful chromosome segregation depends on the ability of sister kinetochores to attach to spindle microtubules. An outer layer of the kinetochore known as the fibrous corona transiently expands in early mitosis and disassembles upon microtubule capture. Neither the functional importance nor the mechanistic basis for this are known. Here we show that the dynein adaptor Spindly and the RZZ kinetochore complex drive fibrous corona formation in a dynein-independent manner. C-terminal farnesylation and MPS1 kinase activity cause conformational changes of Spindly that promote oligomerization of RZZ:Spindly complexes into a corona-like meshwork in cells and in vitro. Concurrent with corona expansion, Spindly potentiates corona shedding by recruiting dynein via three conserved short linear motifs. Expanded, nonsheddable fibrous coronas engage in extensive, long-lived lateral microtubule interactions that persist to metaphase and result in fused sister kinetochores, formation of merotelic attachments and chromosome segregation errors in anaphase.Thus, dynamic kinetochore size regulation in mitosis is coordinated by a single, Spindly-based mechanism that promotes initial microtubule capture and subsequent correct maturation of attachments.

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RZZ and Mad1 dynamics in Drosophila mitosis

Cited by 19 publications

References 45 publications

Polo regulates Spindly to prevent premature stabilization of kinetochore–microtubule attachments

Polo regulates Spindly to prevent premature stabilization of kinetochore–microtubule attachments

Auxin/AID versus conventional knockouts: distinguishing the roles of CENP-T/W in mitotic kinetochore assembly and stability

Dynamic Kinetochore Size Regulation Promotes Microtubule Capture and Chromosome Biorientation in Mitosis

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