Mechanisms of nuclear pore complex disassembly by the mitotic Polo-like kinase 1 (PLK-1) in C. elegans embryos

The nuclear envelope, which protects and organizes the interphase genome, is dismantled during mitosis. In the C. elegans zygote, nuclear envelope breakdown (NEBD) of the parental pronuclei is spatially and temporally regulated during mitosis to promote the unification of the parental genomes. During NEBD, Nuclear Pore Complex (NPC) disassembly is critical for rupturing the nuclear permeability barrier and removing the NPCs from the membranes near the centrosomes and between the juxtaposed pronuclei. By combining live imaging, biochemistry, and phosphoproteomics, we characterized NPC disassembly and unveiled the exact role of the mitotic kinase PLK-1 in this process. We show that PLK-1 disassembles the NPC by targeting multiple NPC sub-complexes, including the cytoplasmic filaments, the central channel, and the inner ring. Notably, PLK-1 is recruited to and phosphorylates intrinsically disordered regions of several multivalent linker nucleoporins, a mechanism that appears to be an evolutionarily conserved driver of NPC disassembly during mitosis. (149/150 words) One-Sentence Summary PLK-1 targets intrinsically disordered regions of multiple multivalent nucleoporins to dismantle the nuclear pore complexes in the C. elegans zygote.

[1]  O. Cohen-Fix,et al.  A membrane reticulum, the centriculum, affects centrosome size and function in Caenorhabditis elegans , 2023, Current Biology.

[2]  E. Dultz,et al.  The Nuclear Pore Complex: Birth, Life, and Death of a Cellular Behemoth , 2022, Cells.

[3]  A. Hoelz,et al.  Architecture of the linker-scaffold in the nuclear pore , 2021, bioRxiv.

[4]  A. Hoelz,et al.  Architecture of the cytoplasmic face of the nuclear pore , 2021, bioRxiv.

[5]  U. Kutay,et al.  Mitotic disassembly and reassembly of nuclear pore complexes. , 2021, Trends in cell biology.

[6]  J. Ellenberg,et al.  A quantitative map of nuclear pore assembly reveals two distinct mechanisms , 2021, bioRxiv.

[7]  Erik E. Griffin,et al.  PLK-1 Regulation of Asymmetric Cell Division in the Early C. elegans Embryo , 2021, Frontiers in Cell and Developmental Biology.

[8]  A. Musacchio,et al.  BUB1 and CENP-U, Primed by CDK1, Are the Main PLK1 Kinetochore Receptors in Mitosis , 2020, Molecular cell.

[9]  M. Dasso,et al.  The Nuclear Pore Complex consists of two independent scaffolds , 2020, bioRxiv.

[10]  L. Van Hove,et al.  PLK-1 promotes the merger of the parental genome into a single nucleus by triggering lamina disassembly , 2020, eLife.

[11]  L. Van Hove,et al.  Phosphorylation of the microtubule-severing AAA+ enzyme Katanin regulates C. elegans embryo development , 2020, The Journal of cell biology.

[12]  O. Cohen-Fix,et al.  C. elegans pronuclei fuse after fertilization through a novel membrane structure , 2019, The Journal of cell biology.

[13]  G. Blobel,et al.  Allosteric modulation of nucleoporin assemblies by intrinsically disordered regions , 2019, Science Advances.

[14]  Fred A. Hamprecht,et al.  ilastik: interactive machine learning for (bio)image analysis , 2019, Nature Methods.

[15]  A. Hoelz,et al.  The Structure of the Nuclear Pore Complex (An Update). , 2019, Annual review of biochemistry.

[16]  Ruedi Aebersold,et al.  A Global Screen for Assembly State Changes of the Mitotic Proteome by SEC-SWATH-MS , 2019, bioRxiv.

[17]  R. Ellis Faculty Opinions recommendation of Robust Genome Editing with Short Single-Stranded and Long, Partially Single-Stranded DNA Donors in Caenorhabditis elegans. , 2019, Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature.

[18]  B. Bowerman,et al.  Mitotic Cell Division in Caenorhabditis elegans , 2019, Genetics.

[19]  J. Cerón,et al.  Efficient Generation of Endogenous Fluorescent Reporters by Nested CRISPR in Caenorhabditis elegans , 2018, Genetics.

[20]  C. Mello,et al.  Robust Genome Editing with Short Single-Stranded and Long, Partially Single-Stranded DNA Donors in Caenorhabditis elegans , 2018, Genetics.

[21]  V. Archambault,et al.  A unified view of spatio-temporal control of mitotic entry: Polo kinase as the key , 2018, Open Biology.

[22]  S. Markert,et al.  Transient and Partial Nuclear Lamina Disruption Promotes Chromosome Movement in Early Meiotic Prophase , 2018, Developmental cell.

[23]  Erik E. Griffin,et al.  Polo-like Kinase Couples Cytoplasmic Protein Gradients in the C. elegans Zygote , 2018, Current Biology.

[24]  J. Ellenberg,et al.  Mechanisms of nuclear pore complex assembly – two different ways of building one molecular machine , 2017, FEBS letters.

[25]  A. Desai,et al.  Channel Nucleoporins Recruit PLK-1 to Nuclear Pore Complexes to Direct Nuclear Envelope Breakdown in C. elegans. , 2017, Developmental cell.

[26]  Wolfram Antonin,et al.  Mitotic Disassembly of Nuclear Pore Complexes Involves CDK1- and PLK1-Mediated Phosphorylation of Key Interconnecting Nucleoporins , 2017, Developmental cell.

[27]  O. Cohen-Fix,et al.  Cell Biology of the Caenorhabditis elegans Nucleus , 2016, Genetics.

[28]  Mark W. Moyle,et al.  A Nucleoporin Docks Protein Phosphatase 1 to Direct Meiotic Chromosome Segregation and Nuclear Assembly. , 2016, Developmental cell.

[29]  W. Wen,et al.  Phospho-Pon Binding-Mediated Fine-Tuning of Plk1 Activity. , 2016, Structure.

[30]  F. Piano,et al.  Identification of Conserved MEL-28/ELYS Domains with Essential Roles in Nuclear Assembly and Chromosome Segregation , 2016, PLoS genetics.

[31]  A. Hoelz,et al.  Architecture of the symmetric core of the nuclear pore , 2016, Science.

[32]  J. Briggs,et al.  Molecular architecture of the inner ring scaffold of the human nuclear pore complex , 2016, Science.

[33]  E. Jorgensen,et al.  SapTrap, a Toolkit for High-Throughput CRISPR/Cas9 Gene Modification in Caenorhabditis elegans , 2016, Genetics.

[34]  Matthias Mann,et al.  Parallel Accumulation-Serial Fragmentation (PASEF): Multiplying Sequencing Speed and Sensitivity by Synchronized Scans in a Trapped Ion Mobility Device. , 2015, Journal of proteome research.

[35]  A. Koide,et al.  Architecture of the fungal nuclear pore inner ring complex , 2015, Science.

[36]  E. Hurt,et al.  Linker Nups connect the nuclear pore complex inner ring with the outer ring and transport channel , 2015, Nature Structural &Molecular Biology.

[37]  M. Gotta,et al.  Cdk1 phosphorylates SPAT-1/Bora to trigger PLK-1 activation and drive mitotic entry in C. elegans embryos , 2015, The Journal of cell biology.

[38]  D. Kachaner,et al.  Understanding the Polo Kinase machine , 2015, Oncogene.

[39]  Joshua A. Arribere,et al.  Efficient Marker-Free Recovery of Custom Genetic Modifications with CRISPR/Cas9 in Caenorhabditis elegans , 2014, Genetics.

[40]  George M. Church,et al.  Heritable genome editing in C. elegans via a CRISPR-Cas9 system , 2013, Nature Methods.

[41]  O. Cohen-Fix,et al.  The dynamic nature of the nuclear envelope , 2013, Nucleus.

[42]  B. Maček,et al.  Dimerization and direct membrane interaction of Nup53 contribute to nuclear pore complex assembly , 2012, The EMBO journal.

[43]  D. Görlich,et al.  The Permeability of Reconstituted Nuclear Pores Provides Direct Evidence for the Selective Phase Model , 2012, Cell.

[44]  Anthony A. Hyman,et al.  A Genome-Scale Resource for In Vivo Tag-Based Protein Function Exploration in C. elegans , 2012, Cell.

[45]  P. Askjaer,et al.  Dissection of the NUP107 nuclear pore subcomplex reveals a novel interaction with spindle assembly checkpoint protein MAD1 in Caenorhabditis elegans , 2012, Molecular biology of the cell.

[46]  K. Oegema,et al.  Affinity purification of protein complexes in C. elegans. , 2011, Methods in cell biology.

[47]  J. Köser,et al.  Distinct association of the nuclear pore protein Nup153 with A- and B-type lamins , 2011, Nucleus.

[48]  U. Kutay,et al.  The nucleoporin Nup88 is interacting with nuclear lamin A , 2011, Molecular biology of the cell.

[49]  Ruedi Aebersold,et al.  Phosphorylation of Nup98 by Multiple Kinases Is Crucial for NPC Disassembly during Mitotic Entry , 2011, Cell.

[50]  Florian Gnad,et al.  PHOSIDA 2011: the posttranslational modification database , 2010, Nucleic Acids Res..

[51]  M. Gotta,et al.  SPAT-1/Bora acts with Polo-like kinase 1 to regulate PAR polarity and cell cycle progression , 2010, Development.

[52]  G. Seydoux,et al.  The C. elegans homolog of nucleoporin Nup98 is required for the integrity and function of germline P granules , 2010, Journal of Cell Science.

[53]  Kyung S. Lee,et al.  Polo-box domain: a versatile mediator of polo-like kinase function , 2010, Cellular and Molecular Life Sciences.

[54]  A. Audhya,et al.  Early embryonic requirement for nucleoporin Nup35/NPP-19 in nuclear assembly. , 2009, Developmental biology.

[55]  Geoffrey J. Barton,et al.  Jalview Version 2—a multiple sequence alignment editor and analysis workbench , 2009, Bioinform..

[56]  R. Wozniak,et al.  Nup53 is required for nuclear envelope and nuclear pore complex assembly. , 2008, Molecular biology of the cell.

[57]  P. Gönczy,et al.  PLK-1 asymmetry contributes to asynchronous cell division of C. elegans embryos , 2008, Development.

[58]  J. Ellenberg,et al.  Systematic kinetic analysis of mitotic dis- and reassembly of the nuclear pore in living cells , 2008, The Journal of cell biology.

[59]  J. Ahringer,et al.  PAR proteins direct asymmetry of the cell cycle regulators Polo-like kinase and Cdc25 , 2008, The Journal of cell biology.

[60]  R. Lin,et al.  Polo kinases regulate C. elegans embryonic polarity via binding to DYRK2-primed MEX-5 and MEX-6 , 2008, Development.

[61]  K. Oegema,et al.  A role for Rab5 in structuring the endoplasmic reticulum , 2007, The Journal of cell biology.

[62]  R. Shiekhattar,et al.  The human Nup107–160 nuclear pore subcomplex contributes to proper kinetochore functions , 2007, The EMBO journal.

[63]  Karen Oegema,et al.  A microtubule-independent role for centrosomes and aurora a in nuclear envelope breakdown. , 2007, Developmental cell.

[64]  M. Hetzer,et al.  MEL‐28/ELYS is required for the recruitment of nucleoporins to chromatin and postmitotic nuclear pore complex assembly , 2007, EMBO reports.

[65]  R. Wozniak,et al.  Vertebrate Nup53 interacts with the nuclear lamina and is required for the assembly of a Nup93-containing complex. , 2005, Molecular biology of the cell.

[66]  Vincent Galy,et al.  Caenorhabditis elegans nucleoporins Nup93 and Nup205 determine the limit of nuclear pore complex size exclusion in vivo. , 2003, Molecular biology of the cell.

[67]  Erich A Nigg,et al.  The crystal structure of the human polo‐like kinase‐1 polo box domain and its phospho‐peptide complex , 2003, The EMBO journal.

[68]  Michael B. Yaffe,et al.  The Molecular Basis for Phosphodependent Substrate Targeting and Regulation of Plks by the Polo-Box Domain , 2003, Cell.

[69]  V. Cordes,et al.  Direct interaction with nup153 mediates binding of Tpr to the periphery of the nuclear pore complex. , 2003, Molecular biology of the cell.

[70]  Michael B Yaffe,et al.  Proteomic Screen Finds pSer/pThr-Binding Domain Localizing Plk1 to Mitotic Substrates , 2003, Science.

[71]  Y. Dong,et al.  Systematic functional analysis of the Caenorhabditis elegans genome using RNAi , 2003, Nature.

[72]  N. Daigle,et al.  An evolutionarily conserved NPC subcomplex, which redistributes in part to kinetochores in mammalian cells , 2001, The Journal of cell biology.

[73]  K Weber,et al.  Essential roles for Caenorhabditis elegans lamin gene in nuclear organization, cell cycle progression, and spatial organization of nuclear pore complexes. , 2000, Molecular biology of the cell.

[74]  K. Wilson,et al.  C. elegans nuclear envelope proteins emerin, MAN1, lamin, and nucleoporins reveal unique timing of nuclear envelope breakdown during mitosis. , 2000, Molecular biology of the cell.

[75]  N. Munakata [Genetics of Caenorhabditis elegans]. , 1989, Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme.

[76]  G. Blobel,et al.  Identification and characterization of a nuclear pore complex protein , 1986, Cell.

[77]  U. Kutay,et al.  Cellular Reorganization during Mitotic Entry. , 2017, Trends in cell biology.

[78]  D. Görlich,et al.  Transport Selectivity of Nuclear Pores, Phase Separation, and Membraneless Organelles. , 2016, Trends in biochemical sciences.

[79]  A. W. E. E. K. L. Y. J. O U R N A L D E V O T E D T O T H E A D V A N C E,et al.  S C I E N C E , 2022 .