SCFSlimb ubiquitin ligase suppresses condensin II–mediated nuclear reorganization by degrading Cap-H2

Buster, Daniel W.; Daniel, Scott G.; Nguyen, Huy; Windler, Sarah L.; Skwarek, Lara C.; Peterson, Maureen; Roberts, Meredith; Meserve, Joy H.; Hartl, Tom A.; Klebba, Joseph E.; Bilder, David; Bosco, Giovanni; Rogers, Gerald S.

doi:10.1083/jcb.201207183

Cited by 73 publications

(153 citation statements)

References 55 publications

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“…GFP-binding protein (GBP) (53) was fused to the Fc domain of human IgG, tagged with His 6 in pET28a (EMD Biosciences), expressed in Escherichia coli, and purified on HisPur resin (Thermo Fisher) according to the manufacturer's instructions (54). Purified GBP was bound to Protein A-coupled Sepharose, then was cross-linked to the resin by incubation with 20 mM dimethyl pimelimidate dihydrochloride in PBS, pH 8.3, for 2 h at 22°C; the coupling reaction was quenched by incubation with 0.2 M ethanolamine, pH 8.3, for 1 h at 22°C.…”

Section: Methodsmentioning

confidence: 99%

“…GBP-coated beads were rocked with lysate for 1 h at 4°C, washed two times with 1 mL CLB, and then boiled in Laemmli sample buffer. In vivo ubiquitination assays were performed by coexpressing Plk4-EGFP constructs with triple FLAG-tagged Drosophila ubiquitin (CG32744) (also under the metallothionein promoter and Cuinduced) (54) and then probing the immunoblot of the cell lysate with anti-FLAG antibody (Sigma).…”

Section: Methodsmentioning

confidence: 99%

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Autoinhibition and relief mechanism for Polo-like kinase 4

Klebba

Buster

McLamarrah

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Polo-like kinase 4 (Plk4) is a master regulator of centriole duplication, and its hyperactivity induces centriole amplification. Homodimeric Plk4 has been shown to be ubiquitinated as a result of autophosphorylation, thus promoting its own degradation and preventing centriole amplification. Unlike other Plks, Plk4 contains three rather than two Polo box domains, and the function of its third Polo box (PB3) is unclear. Here, we performed a functional analysis of Plk4's structural domains. Like other Plks, Plk4 possesses a previously unidentified autoinhibitory mechanism mediated by a linker (L1) near the kinase domain. Thus, autoinhibition is a conserved feature of Plks. In the case of Plk4, autoinhibition is relieved after homodimerization and is accomplished by PB3 and by autophosphorylation of L1. In contrast, autophosphorylation of the second linker promotes separation of the Plk4 homodimer. Therefore, autoinhibition delays the multiple consequences of activation until Plk4 dimerizes. These findings reveal a complex mechanism of Plk4 regulation and activation which govern the process of centriole duplication.autoinhibition | centriole | centrosome | Polo box | Polo-like kinase 4

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Autoinhibition and relief mechanism for Polo-like kinase 4

Klebba

Buster

McLamarrah

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…Taken all together, it is reasonable to speculate that condensin II regulates not only the resolution of sister chromatids within individual chromosomes, but also the disruption of interchromosomal interactions within the interphase nucleus. Evidence has also been provided that such interphase functions of condensin II are down-regulated by the SCF slimb ubiquitin ligase complex (Buster et al 2013). It remains to be clarified, however, whether Drosophila indeed has a conventional type of condensin II because the Cap-G2 gene is apparently missing in its genome (Herzog et al 2013).…”

Section: Condensin II Initiates Its Action Long Before Entry Into Mitmentioning

confidence: 99%

Chromosome Dynamics during Mitosis

Hirano

2015

Cold Spring Harb Perspect Biol

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The primary goal of mitosis is to partition duplicated chromosomes into daughter cells. Eukaryotic chromosomes are equipped with two distinct classes of intrinsic machineries, cohesin and condensins, that ensure their faithful segregation during mitosis. Cohesin holds sister chromatids together immediately after their synthesis during S phase until the establishment of bipolar attachments to the mitotic spindle in metaphase. Condensins, on the other hand, attempt to "resolve" sister chromatids by counteracting cohesin. The products of the balancing acts of cohesin and condensins are metaphase chromosomes, in which two rod-shaped chromatids are connected primarily at the centromere. In anaphase, this connection is released by the action of separase that proteolytically cleaves the remaining population of cohesin. Recent studies uncover how this series of events might be mechanistically coupled with each other and intricately regulated by a number of regulatory factors.

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“…In contrast, Slimb has been identified as a pairing-promoting factor that negatively regulates the Cap-H2 anti-pairing activity [18,20]. Therefore, we predicted that CK1α also is a pairing promoting factor not identified in previous chromosome pairing screens.…”

Section: Resultsmentioning

confidence: 88%

Drosophila Casein Kinase I Alpha Regulates Homolog Pairing and Genome Organization by Modulating Condensin II Subunit Cap-H2 Levels

et al. 2015

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The spatial organization of chromosomes within interphase nuclei is important for gene expression and epigenetic inheritance. Although the extent of physical interaction between chromosomes and their degree of compaction varies during development and between different cell-types, it is unclear how regulation of chromosome interactions and compaction relate to spatial organization of genomes. Drosophila is an excellent model system for studying chromosomal interactions including homolog pairing. Recent work has shown that condensin II governs both interphase chromosome compaction and homolog pairing and condensin II activity is controlled by the turnover of its regulatory subunit Cap-H2. Specifically, Cap-H2 is a target of the SCFSlimb E3 ubiquitin-ligase which down-regulates Cap-H2 in order to maintain homologous chromosome pairing, chromosome length and proper nuclear organization. Here, we identify Casein Kinase I alpha (CK1α) as an additional negative-regulator of Cap-H2. CK1α-depletion stabilizes Cap-H2 protein and results in an accumulation of Cap-H2 on chromosomes. Similar to Slimb mutation, CK1α depletion in cultured cells, larval salivary gland, and nurse cells results in several condensin II-dependent phenotypes including dispersal of centromeres, interphase chromosome compaction, and chromosome unpairing. Moreover, CK1α loss-of-function mutations dominantly suppress condensin II mutant phenotypes in vivo. Thus, CK1α facilitates Cap-H2 destruction and modulates nuclear organization by attenuating chromatin localized Cap-H2 protein.

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SCFSlimb ubiquitin ligase suppresses condensin II–mediated nuclear reorganization by degrading Cap-H2

Abstract: SCFSlimb-mediated down-regulation of the condensin II subunit Cap-H2 is required to maintain proper organization and morphology of the interphase nucleus.

Cited by 73 publications

References 55 publications

Autoinhibition and relief mechanism for Polo-like kinase 4

Autoinhibition and relief mechanism for Polo-like kinase 4

Chromosome Dynamics during Mitosis

Drosophila Casein Kinase I Alpha Regulates Homolog Pairing and Genome Organization by Modulating Condensin II Subunit Cap-H2 Levels

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