title: Downregulation of protein 4.1R, a mature centriole protein, disrupts centrosomes, alters cell cycle progression, and perturbs mitotic spindles and anaphase.
abstract: Centrosomes nucleate and organize interphase microtubules and are instrumental in mitotic bipolar spindle assembly, ensuring orderly cell cycle progression with accurate chromosome segregation. We report that the multifunctional structural protein 4.1R localizes at centrosomes to distal/subdistal regions of mature centrioles in a cell cycle-dependent pattern. Significantly, 4.1R-specific depletion mediated by RNA interference perturbs subdistal appendage proteins ninein and outer dense fiber 2/cenexin at mature centrosomes and concomitantly reduces interphase microtubule anchoring and organization. 4.1R depletion causes G(1) accumulation in p53-proficient cells, similar to depletion of many other proteins that compromise centrosome integrity. In p53-deficient cells, 4.1R depletion delays S phase, but aberrant ninein distribution is not dependent on the S-phase delay. In 4.1R-depleted mitotic cells, efficient centrosome separation is reduced, resulting in monopolar spindle formation. Multipolar spindles and bipolar spindles with misaligned chromatin are also induced by 4.1R depletion. Notably, all types of defective spindles have mislocalized NuMA (nuclear mitotic apparatus protein), a 4.1R binding partner essential for spindle pole focusing. These disruptions contribute to lagging chromosomes and aberrant microtubule bridges during anaphase/telophase. Our data provide functional evidence that 4.1R makes crucial contributions to the structural integrity of centrosomes and mitotic spindles which normally enable mitosis and anaphase to proceed with the coordinated precision required to avoid pathological events.
authors.affiliation: Department of Cell Biology and Imaging, University of California-LBNL, 1 Cyclotron Road, MS 74-157, Berkeley, CA 94720, USA.
authors.forename: Sharon Wald
authors.initials: SW
authors.lastname: Krauss
authors.forename: Jeffrey R
authors.initials: JR
authors.lastname: Spence
authors.forename: Shirin
authors.initials: S
authors.lastname: Bahmanyar
authors.forename: Angela I M
authors.initials: AI
authors.lastname: Barth
authors.forename: Minjoung M
authors.initials: MM
authors.lastname: Go
authors.forename: Debra
authors.initials: D
authors.lastname: Czerwinski
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Meyer
mesh_terms: D000705:Anaphase / Q000502:physiology; D034961:Antigens, Nuclear / Q000032:analysis; D002453:Cell Cycle / Q000502:physiology*; D018797:Cell Cycle Proteins; D002460:Cell Line / Q000648:ultrastructure; D002502:Centrioles / Q000737:chemistry*; D018385:Centrosome / Q000378:metabolism / Q000648:ultrastructure*; D020090:Chromosome Segregation / Q000502:physiology; D003598:Cytoskeletal Proteins / Q000032:analysis / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D015536:Down-Regulation; D006367:HeLa Cells / Q000648:ultrastructure; D006801:Humans; D007399:Interphase / Q000502:physiology; D008565:Membrane Proteins / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D008870:Microtubules / Q000648:ultrastructure; D034521:Nuclear Matrix-Associated Proteins / Q000032:analysis; D009687:Nuclear Proteins / Q000032:analysis; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000494:pharmacology; D008941:Spindle Apparatus / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure*; D016159:Tumor Suppressor Protein p53 / Q000502:physiology
chemical_list: D034961:Antigens, Nuclear; D018797:Cell Cycle Proteins; D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; C512806:NIN protein, human; C050045:NUMA1 protein, human; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D016159:Tumor Suppressor Protein p53; C028841:erythrocyte membrane band 4.1 protein