Downregulation of Protein 4.1R, a Mature Centriole Protein, Disrupts Centrosomes, Alters Cell Cycle Progression, and Perturbs Mitotic Spindles and Anaphase

Krauss, Sharon Wald; Spence, Jeffrey R.; Bahmanyar, Shirin; Barth, Angela I.M.; Go, Minjoung M.; Czerwinski, Debra K.; Meyer, Adam

doi:10.1128/mcb.02021-07

Cited by 32 publications

(31 citation statements)

References 75 publications

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“…Moreover, MTs growing close to the cortex in 4.1R-KD cells undergo fewer episodes of catastrophe, longer periods of growth and less time pausing and shortening, compared with control MTs. Although a defective MT nucleation from the centrosome could account for some of these effects, no reduction in MT nucleation has been observed in 4.1R-KD cells (Krauss et al, 2008). We therefore assume that the effects on the dynamic behavior of the MT network at the cell edge that we describe here are specifically due to a lack of 4.1R at the cell cortex.…”

Section: Discussionmentioning

confidence: 86%

Protein 4.1R binds to CLASP2 and regulates dynamics, organization and attachment of microtubules to the cell cortex

Ruiz-Sáenz¹,

Haren²,

Sayas³

et al. 2013

Journal of Cell Science

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SummaryThe microtubule (MT) cytoskeleton is essential for many cellular processes, including cell polarity and migration. Cortical platforms, formed by a subset of MT plus-end-tracking proteins, such as CLASP2, and non-MT binding proteins such as LL5b, attach distal ends of MTs to the cell cortex. However, the mechanisms involved in organizing these platforms have not yet been described in detail. Here we show that 4.1R, a FERM-domain-containing protein, interacts and colocalizes with cortical CLASP2 and is required for the correct number and dynamics of CLASP2 cortical platforms. Protein 4.1R also controls binding of CLASP2 to MTs at the cell edge by locally altering GSK3 activity. Furthermore, in 4.1R-knockdown cells MT plus-ends were maintained for longer in the vicinity of cell edges, but instead of being tethered to the cell cortex, MTs continued to grow, bending at cell margins and losing their radial distribution. Our results suggest a previously unidentified role for the scaffolding protein 4.1R in locally controlling CLASP2 behavior, CLASP2 cortical platform turnover and GSK3 activity, enabling correct MT organization and dynamics essential for cell polarity.

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Section: Discussionmentioning

confidence: 86%

Protein 4.1R binds to CLASP2 and regulates dynamics, organization and attachment of microtubules to the cell cortex

Ruiz-Sáenz¹,

Haren²,

Sayas³

et al. 2013

Journal of Cell Science

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“…Nucleolin could anchor microtubules directly or via other anchoring protein localizing at the mature centriole such as ninein 14 since nucleolin antibodies immunoprecipitated ninein, or through p150 glued , APC, FOP, CAP350 and EB1, [40][41][42] Kif3a, 43 nudel 44 or 4.1R-135. 45 Previous studies have shown the presence of extra centrosomes markers containing structure in mitotic cells silenced for nucleolin. 28,29 Thus, it was suggested that nucleolin may act as a negative regulator of centriole duplication as it is the case for B23, 46 p53, 47 cdk1, 48 BRCA1 49 and CDK5RAP2.…”

Section: Discussionmentioning

confidence: 99%

Centrosomal nucleolin is required for microtubule network organization

et al. 2015

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Nucleolin is a pleiotropic protein involved in a variety of cellular processes. Although multipolar spindle formation has been observed after nucleolin depletion, the roles of nucleolin in centrosome regulation and functions have not been addressed. Here we report using immunofluorescence and biochemically purified centrosomes that nucleolin colocalized only with one of the centrioles during interphase which was further identified as the mature centriole. Upon nucleolin depletion, cells exhibited an amplification of immature centriole markers surrounded by irregular pericentrin staining; these structures were exempt from maturation markers and unable to nucleate microtubules. Furthermore, the microtubule network was disorganized in these cells, exhibiting frequent non-centrosomal microtubules. At the mature centriole a reduced kinetics in the centrosomal microtubule nucleation phase was observed in live silenced cells, as well as a perturbation of microtubule anchoring. Immunoprecipitation experiments showed that nucleolin belongs to protein complexes containing 2 key centrosomal proteins, g-tubulin and ninein, involved in microtubule nucleation and anchoring steps. Altogether, our study uncovered a new role for nucleolin in restricting microtubule nucleation and anchoring at centrosomes in interphase cells.

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“…We previously reported that 4.1R depletion causes microtubule disorganization in vitro and in intact cells (Krauss et al, 2002;Krauss et al, 2004;Krauss et al, 2008). We reasoned that 4.1R could have significant interactions at the nucleo-cytoskeletal interface affecting centrosome tethering and microtubules, particularly if 4.1R were functionally associated with emerin and lamin A.…”

Section: Increased Centrosome-nuclear Envelope Distance After 41r Dementioning

confidence: 99%

“…It was subsequently discovered, however, that nucleated cells express multiple 4.1R isoforms at several subcellular locations (De Carcer et al, 1995;Delhommeau et al, 2002;Krauss et al, 1997b;Krauss et al, 1997a). In nucleated cells, protein 4.1R not only organizes plasma-membrane-cytoskeletal microstructure, but, as we have previously shown, it is also integral to mitotic spindle and centrosome assembly and structure (Krauss et al, 2004;Krauss et al, 2008). Furthermore, we found that 4.1R epitopes in mammalian and Xenopus laevis nuclei are partially coincident with RNA splicing and/or transcription factories, DNA replication assemblies, nuclear pores and the nuclear periphery, and are largely coincident with nuclear anti-actin signals in nuclear matrix (Krauss et al, 2003;Krauss et al, 1997a).…”

Section: Introductionmentioning

confidence: 99%

“…If 4.1R has a functional association with emerin, 4.1R depletion might also alter nuclear morphology. To analyze the effects of 4.1R depletion on nuclear morphology in intact mammalian cells, we examined DAPI-stained cells after 4.1R downregulation by RNAi (Krauss et al, 2008) and MEFs from 4.1R AUG1,2-/-mice relative to matched MEFs from wild-type mice. We found that 4.1R-deficient cells have an ~sixfold increase in perturbed nuclei (those that are markedly irregular with blebbing 1436 Journal of Cell Science 124 (9) (C)Localization of nuclear envelope proteins by indirect immunofluorescence.…”

Section: 1r Depletion Induces Nuclear Dysmorphology With Altered Emmentioning

confidence: 99%

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Structural protein 4.1R is integrally involved in nuclear envelope protein localization, centrosome–nucleus association and transcriptional signaling

Meyer

Almendrala

et al. 2011

Journal of Cell Science

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SummaryThe multifunctional structural protein 4.1R is required for assembly and maintenance of functional nuclei but its nuclear roles are unidentified. 4.1R localizes within nuclei, at the nuclear envelope, and in cytoplasm. Here we show that 4.1R, the nuclear envelope protein emerin and the intermediate filament protein lamin A/C co-immunoprecipitate, and that 4.1R-specific depletion in human cells by RNA interference produces nuclear dysmorphology and selective mislocalization of proteins from several nuclear subcompartments. Such 4.1R-deficiency causes emerin to partially redistribute into the cytoplasm, whereas lamin A/C is disorganized at nuclear rims and displaced from nucleoplasmic foci. The nuclear envelope protein MAN1, nuclear pore proteins Tpr and Nup62, and nucleoplasmic proteins NuMA and LAP2 also have aberrant distributions, but lamin B and LAP2 have normal localizations. 4.1R-deficient mouse embryonic fibroblasts show a similar phenotype. We determined the functional effects of 4.1R-deficiency that reflect disruption of the association of 4.1R with emerin and A-type lamin: increased nucleus-centrosome distances, increased -catenin signaling, and relocalization of -catenin from the plasma membrane to the nucleus. Furthermore, emerin-and lamin-A/C-null cells have decreased nuclear 4.1R. Our data provide evidence that 4.1R has important functional interactions with emerin and A-type lamin that impact upon nuclear architecture, centrosome-nuclear envelope association and the regulation of -catenin transcriptional co-activator activity that is dependent on -catenin nuclear export.

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Downregulation of Protein 4.1R, a Mature Centriole Protein, Disrupts Centrosomes, Alters Cell Cycle Progression, and Perturbs Mitotic Spindles and Anaphase

Cited by 32 publications

References 75 publications

Protein 4.1R binds to CLASP2 and regulates dynamics, organization and attachment of microtubules to the cell cortex

Protein 4.1R binds to CLASP2 and regulates dynamics, organization and attachment of microtubules to the cell cortex

Centrosomal nucleolin is required for microtubule network organization

Structural protein 4.1R is integrally involved in nuclear envelope protein localization, centrosome–nucleus association and transcriptional signaling

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