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Mol. Cell. Biol. doi:10.1128/MCB.02021-07
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Downregulation of protein 4.1R, a mature centriole protein, disrupts centrosomes, alters cell cycle progression, and perturbs mitotic spindles and anaphase

Department of Cell Biology and Imaging, University of California-Lawrence Berkeley National Laboratory, Berkeley, CA 94720, Departments of Molecular and Cellular Physiology and Biological Sciences, Stanford University, Stanford CA 94305-5435, Department of Medicine, Stanford University School of Medicine, Stanford CA 94305-5435

^* To whom correspondence should be addressed. Email: sakrauss{at}lbl.gov.

	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 ODF2/cenexin at mature centrosomes and concomitantly reduces interphase microtubule anchoring and organization. 4.1R-depletion causes G1 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 centrosome and mitotic spindle structural integrity which normally enable mitosis and anaphase to proceed with the coordinated precision required to avoid pathological events.