CDK1 controls CHMP7-dependent nuclear envelope reformation

Through membrane sealing and disassembly of spindle microtubules, the Endosomal Sorting Complex Required for Transport-III (ESCRT-III) machinery has emerged as a key player in the regeneration of a sealed nuclear envelope (NE) during mitotic exit, and in the repair of this organelle during interphase rupture. ESCRT-III assembly at the NE occurs transiently during mitotic (M) exit and is initiated when CHMP7, an ER-localised ESCRT-II/ESCRT-III hybrid protein, interacts with the Inner Nuclear Membrane (INM) protein LEM2. Whilst classical nucleocytoplasmic transport mechanisms have been proposed to separate LEM2 and CHMP7 during interphase, it is unclear how CHMP7 assembly is suppressed in mitosis when NE and ER identities are mixed. Here, we use live cell imaging and protein biochemistry to examine the biology of these proteins during M-exit. Firstly, we show that CHMP7 plays an important role in the dissolution of LEM2 clusters that form at the NE during M-exit. Secondly, we show that CDK1 phosphorylates CHMP7 upon M-entry at Ser3 and Ser441 and that this phosphorylation reduces CHMP7’s interaction with LEM2, limiting its assembly during M-phase. We show that spatiotemporal differences in the dephosphorylation of CHMP7 license its assembly at the NE during telophase, but restrict its assembly on the ER at this time. Without CDK1 phosphorylation, CHMP7 undergoes inappropriate assembly in the peripheral ER during M-exit, capturing LEM2 and downstream ESCRT-III components. Lastly, we establish that a microtubule network is dispensable for ESCRT-III assembly at the reforming nuclear envelope. These data identify a key cell-cycle control programme allowing ESCRT-III-dependent nuclear regeneration

Estudo morfológico de superfícies ósseas após secção por pontas diamantadas ou laser de érbio: YAG Morphological evaluation of bone surfaces after sectioning with diamond points or erbium: YAG laser

O objetivo deste trabalho foi analisar morfologicamente as superfícies ósseas resultantes da secção por pontas diamantadas ou por laser de érbio:YAG. Cinco ratos (Rattus norvegicus albinus foram sacrificados por dose letal de fenobarbital. Após a execução deste procedimento, os ossos predeterminados foram submetidos à secção por pontas diamantadas ou por laser de érbio:YAG em uma energia de 300 mJ por pulso e taxa de repetição de 2 Hz. As amostras foram submetidas a análise em microscópio eletrônico de varredura, revelando a existência de um padrão para as secções obtidas com cada instrumento, sendo verificada uma superfície mais regular nas amostras seccionadas com o laser de érbio:YAG. Em aumentos da ordem de 3000 vezes, pode-se observar indícios de fusão e seqüente solidificação das superfícies seccionadas por meio do laser de érbio:YAG. Conclui-se que o laser de érbio:YAG foi eficaz na remoção de tecido ósseo, mas que, nos parâmetros utilizados neste estudo, foi responsável por alterações morfológicas sugestivas de significativo aumento de temperatura, não devendo ser indicado, nestas condições, para a execução de secções ósseas.<br>The purpose of this study was to analyze bone surfaces cut by Er:YAG laser or by diamond points. Five male rats (Rattus norvegicus albinus) were killed by lethal dose of phenobarbital, and selected bones were cut by Er:YAG laser at 300 mJ and 2 Hz, or by diamond points. The samples were submitted to evaluation through SEM at standard magnifications. By analyzing the surfaces, the authors could observe a pattern for each group, with a smoother surface in the laser cut group than in the diamond cut group. In higher magnifications, the images suggest melting and solidifying in the laser group. The authors conclude that the laser was effective in bone cutting. However, it was responsible for morphological changes that suggest a significant temperature increase

AKTIP interacts with ESCRT I and is needed for the recruitment of ESCRT III subunits to the midbody

Funding: This work was supported by PRF 2016-67, Progetti di Ricerca, Sapienza University of Rome (RP1181642E87148C), AIRC IG-24614 to IS, FIRC (22392) to MLT and IS, CIB (http://www.cibiotech.it/) and Fondazione Buzzati Traverso (813 - https://www.fondazioneadrianobuzzatitraverso.it) to MLT and IS; Avvio alla Ricerca, Sapienza University of Rome (AR2181642B6F2E48, AR1181642EE61111) to RB, SDG and IS. CM has been supported by EMBO ST fellowship 7621, Veronesi TG 2019. MLT is supported by Be For ERC, Sapienza. IC is supported by R01GM117376 and NSF Career 1751197. JGC is supported by a Wellcome Trust Senior Research Fellowship 206346/Z/17/Z. This work was supported in part by the Francis Crick Institute which receives its core funding from Cancer Research UK (FC001002), the UK Medical Research Council (FC001002), and the Wellcome Trust (FC001002). For the purpose of Open Access, the author has applied a CC BY public copyright licence to any Author Accepted Manuscript version arising from this submission. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.To complete mitosis, the bridge that links the two daughter cells needs to be cleaved. This step is carried out by the endosomal sorting complex required for transport (ESCRT) machinery. AKTIP, a protein discovered to be associated with telomeres and the nuclear membrane in interphase cells, shares sequence similarities with the ESCRT I component TSG101. Here we present evidence that during mitosis AKTIP is part of the ESCRT machinery at the midbody. AKTIP interacts with the ESCRT I subunit VPS28 and forms a circular supra-structure at the midbody, in close proximity with TSG101 and VPS28 and adjacent to the members of the ESCRT III module CHMP2A, CHMP4B and IST1. Mechanistically, the recruitment of AKTIP is dependent on MKLP1 and independent of CEP55. AKTIP and TSG101 are needed together for the recruitment of the ESCRT III subunit CHMP4B and in parallel for the recruitment of IST1. Alone, the reduction of AKTIP impinges on IST1 and causes multinucleation. Our data altogether reveal that AKTIP is a component of the ESCRT I module and functions in the recruitment of ESCRT III components required for abscission.Depto. de Biología CelularFac. de Ciencias BiológicasTRUEpu