Cell cycle progression by the repression of primary cilia formation in proliferating cells

We have isolated the murine homologue of the human Ki-67 antigen. The Ki-67 antigen is used as a marker to assess the proliferative capacity of tumour cells; however, its cellular function is not known. The murine Ki-67 cDNA sequence (TSG126) was found to contain 13 tandem repeats, making up more than half of the total protein size. A comparison of this repetitive sequence block to its human counterpart, which contains 16 consecutive repeat units, revealed several conserved sequence motifs, including one motif frequently observed in proteins interacting with DNA. An antiserum developed against the product of the TSG126 cDNA clone identified a protein with an apparent molecular mass of 360 kDa, mainly expressed in proliferating cells. The TSG126 protein begins to accumulate during the late G1 stage of the cell cycle and is first seen as numerous small granules evenly distributed throughout the nucleus. During the S and the G2 phases, larger foci that overlap with the nucleoli and the heterochromatic regions are formed. At the onset of mitosis the TSG126 protein undergoes a dramatic redistribution process and becomes associated with the surface of the condensed chromosomes. The relative absence of the TSG126 protein from G1 interphase cells strongly argues against a model where the association of the TSG126 protein with mitotic chromosomes merely reflects a mechanism for the symmetrical distribution of nucleolar proteins between daughter cells. Instead, the intracellular distribution of the TSG126 protein during the cell cycle suggests that it could have a chromatin-associated function in both interphase and mitotic cells. Microinjection of anti-TSG126 antibodies into proliferating Swiss-3T3 fibroblasts was found to delay cell cycle progression, indicating that the TSG126 protein has an essential nuclear function.

Download Full-text

Deregulated expression of the RanBP1 gene alters cell cycle progression in murine fibroblasts

Journal of Cell Science ◽

10.1242/jcs.110.19.2345 ◽

1997 ◽

Vol 110 (19) ◽

pp. 2345-2357 ◽

Cited By ~ 1

Author(s):

A. Battistoni ◽

G. Guarguaglini ◽

F. Degrassi ◽

C. Pittoggi ◽

A. Palena ◽

...

Keyword(s):

Cell Cycle ◽

Dna Replication ◽

Cell Cycle Progression ◽

Protein Import ◽

Proliferating Cells ◽

Cycle Progression ◽

Ran Gtpase ◽

Genes Encoding ◽

Active Transcription ◽

Gtpase Cycle

RanBP1 is a molecular partner of the Ran GTPase, which is implicated in the control of several processes, including DNA replication, mitotic entry and exit, cell cycle progression, nuclear structure, protein import and RNA export. While most genes encoding Ran-interacting partners are constitutively active, transcription of the RanBP1 mRNA is repressed in non proliferating cells, is activated at the G1/S transition in cycling cells and peaks during S phase. We report here that forced expression of the RanBP1 gene disrupts the orderly execution of the cell division cycle at several stages, causing inhibition of DNA replication, defective mitotic exit and failure of chromatin decondensation during the telophase-to-interphase transition in cells that achieve nuclear duplication and chromosome segregation. These results suggest that deregulated RanBP1 activity interferes with the Ran GTPase cycle and prevents the functioning of the Ran signalling system during the cell cycle.

Download Full-text

KIF14 controls ciliogenesis via regulation of Aurora A and is important for Hedgehog signaling

The Journal of Cell Biology ◽

10.1083/jcb.201904107 ◽

2020 ◽

Vol 219 (6) ◽

Cited By ~ 6

Author(s):

Petra Pejskova ◽

Madeline Louise Reilly ◽

Lucia Bino ◽

Ondrej Bernatik ◽

Linda Dolanska ◽

...

Keyword(s):

Cell Cycle ◽

Primary Cilium ◽

Hedgehog Signaling ◽

Cell Cycle Progression ◽

Primary Cilia ◽

Aurora A ◽

Pathway Activation ◽

Cilia Formation ◽

Tightly Coupled ◽

Hh Signaling

Primary cilia play critical roles in development and disease. Their assembly and disassembly are tightly coupled to cell cycle progression. Here, we present data identifying KIF14 as a regulator of cilia formation and Hedgehog (HH) signaling. We show that RNAi depletion of KIF14 specifically leads to defects in ciliogenesis and basal body (BB) biogenesis, as its absence hampers the efficiency of primary cilium formation and the dynamics of primary cilium elongation, and disrupts the localization of the distal appendage proteins SCLT1 and FBF1 and components of the IFT-B complex. We identify deregulated Aurora A activity as a mechanism contributing to the primary cilium and BB formation defects seen after KIF14 depletion. In addition, we show that primary cilia in KIF14-depleted cells are defective in response to HH pathway activation, independently of the effects of Aurora A. In sum, our data point to KIF14 as a critical node connecting cell cycle machinery, effective ciliogenesis, and HH signaling.

Download Full-text

Plk4 triggers autonomous de novo centriole biogenesis and maturation

The Journal of Cell Biology ◽

10.1083/jcb.202008090 ◽

2021 ◽

Vol 220 (5) ◽

Author(s):

Catarina Nabais ◽

Delphine Pessoa ◽

Jorge de-Carvalho ◽

Thomas van Zanten ◽

Paulo Duarte ◽

...

Keyword(s):

Cell Cycle ◽

Cell Cycle Progression ◽

De Novo ◽

Cell Types ◽

Proliferating Cells ◽

Cycle Progression ◽

Controlled System ◽

Pericentriolar Material ◽

Centriole Assembly ◽

Critical Components

Centrioles form centrosomes and cilia. In most proliferating cells, centrioles assemble through canonical duplication, which is spatially, temporally, and numerically regulated by the cell cycle and the presence of mature centrioles. However, in certain cell types, centrioles assemble de novo, yet by poorly understood mechanisms. Herein, we established a controlled system to investigate de novo centriole biogenesis, using Drosophila melanogaster egg explants overexpressing Polo-like kinase 4 (Plk4), a trigger for centriole biogenesis. We show that at a high Plk4 concentration, centrioles form de novo, mature, and duplicate, independently of cell cycle progression and of the presence of other centrioles. Plk4 concentration determines the temporal onset of centriole assembly. Moreover, our results suggest that distinct biochemical kinetics regulate de novo and canonical biogenesis. Finally, we investigated which other factors modulate de novo centriole assembly and found that proteins of the pericentriolar material (PCM), and in particular γ-tubulin, promote biogenesis, likely by locally concentrating critical components.

Download Full-text

Nek5 promotes centrosome integrity in interphase and loss of centrosome cohesion in mitosis

The Journal of Cell Biology ◽

10.1083/jcb.201412099 ◽

2015 ◽

Vol 209 (3) ◽

pp. 339-348 ◽

Cited By ~ 24

Author(s):

Suzanna L. Prosser ◽

Navdeep K. Sahota ◽

Laurence Pelletier ◽

Ciaran G. Morrison ◽

Andrew M. Fry

Keyword(s):

Cell Cycle ◽

Chromosome Segregation ◽

Cell Cycle Progression ◽

Primary Cilia ◽

Microtubule Nucleation ◽

Cycle Progression ◽

Spindle Formation ◽

Mitotic Entry ◽

Bipolar Spindle ◽

Centrosome Separation

Nek5 is a poorly characterized member of the NIMA-related kinase family, other members of which play roles in cell cycle progression and primary cilia function. Here, we show that Nek5, similar to Nek2, localizes to the proximal ends of centrioles. Depletion of Nek5 or overexpression of kinase-inactive Nek5 caused unscheduled separation of centrosomes in interphase, a phenotype also observed upon overexpression of active Nek2. However, separated centrosomes that resulted from Nek5 depletion remained relatively close together, exhibited excess recruitment of the centrosome linker protein rootletin, and had reduced levels of Nek2. In addition, Nek5 depletion led to loss of PCM components, including γ-tubulin, pericentrin, and Cdk5Rap2, with centrosomes exhibiting reduced microtubule nucleation. Upon mitotic entry, Nek5-depleted cells inappropriately retained centrosome linker components and exhibited delayed centrosome separation and defective chromosome segregation. Hence, Nek5 is required for the loss of centrosome linker proteins and enhanced microtubule nucleation that lead to timely centrosome separation and bipolar spindle formation in mitosis.

Download Full-text

HDAC6 Signaling at Primary Cilia Promotes Proliferation and Restricts Differentiation of Glioma Cells

Cancers ◽

10.3390/cancers13071644 ◽

2021 ◽

Vol 13 (7) ◽

pp. 1644

Author(s):

Ping Shi ◽

Lan B. Hoang-Minh ◽

Jia Tian ◽

Alice Cheng ◽

Reemsha Basrai ◽

...

Keyword(s):

Tumor Cell ◽

Cell Cycle Progression ◽

Primary Cilia ◽

Glioma Cells ◽

Histone Deacetylase 6 ◽

Cycle Progression ◽

Alpha Tubulin ◽

Antiproliferative Effects ◽

Low Concentrations ◽

Cilia Formation

Histone deacetylase 6 (HDAC6) is an emerging therapeutic target that is overexpressed in glioblastoma when compared to other HDACs. HDAC6 catalyzes the deacetylation of alpha-tubulin and mediates the disassembly of primary cilia, a process required for cell cycle progression. HDAC6 inhibition disrupts glioma proliferation, but whether this effect is dependent on tumor cell primary cilia is unknown. We found that HDAC6 inhibitors ACY-1215 (1215) and ACY-738 (738) inhibited the proliferation of multiple patient-derived and mouse glioma cells. While both inhibitors triggered rapid increases in acetylated alpha-tubulin (aaTub) in the cytosol and led to increased frequencies of primary cilia, they unexpectedly reduced the levels of aaTub in the cilia. To test whether the antiproliferative effects of HDAC6 inhibitors are dependent on tumor cell cilia, we generated patient-derived glioma lines devoid of cilia through depletion of ciliogenesis genes ARL13B or KIF3A. At low concentrations, 1215 or 738 did not decrease the proliferation of cilia-depleted cells. Moreover, the differentiation of glioma cells that was induced by HDAC6 inhibition did not occur after the inhibition of cilia formation. These data suggest HDAC6 signaling at primary cilia promotes the proliferation of glioma cells by restricting their ability to differentiate. Surprisingly, overexpressing HDAC6 did not reduce cilia length or the frequency of ciliated glioma cells, suggesting other factors are required to control HDAC6-mediated cilia disassembly in glioma cells. Collectively, our findings suggest that HDAC6 promotes the proliferation of glioma cells through primary cilia.

Download Full-text

Plk4 triggers autonomous de novo centriole biogenesis and maturation

10.1101/2020.04.29.068650 ◽

2020 ◽

Author(s):

Catarina Nabais ◽

Delphine Pessoa ◽

Jorge de-Carvalho ◽

Thomas van Zanten ◽

Paulo Duarte ◽

...

Keyword(s):

Cell Cycle ◽

Cell Cycle Progression ◽

De Novo ◽

Cell Types ◽

Proliferating Cells ◽

Cycle Progression ◽

Controlled System ◽

Centriole Assembly ◽

Critical Components ◽

Kinetics Of

AbstractCentrioles form centrosomes and cilia. In most proliferating cells, centrioles assemble through canonical duplication, which is spatially, temporally and numerically regulated by the cell cycle and the presence of mature centrioles. However, in certain cell-types, centrioles assemble de novo, yet by poorly understood mechanisms. Here, we established a controlled system to investigate de novo centriole biogenesis, using Drosophila melanogaster egg explants overexpressing Polo-like kinase 4 (Plk4), a trigger for centriole biogenesis. We show that at high Plk4 concentration, centrioles form de novo, mature and duplicate, independently of cell cycle progression and of the presence of other centrioles. Plk4 concentration determines the kinetics of centriole assembly. Moreover, our results suggest Plk4 operates in a switch-like manner to control the onset of de novo centriole formation, and that distinct biochemical kinetics regulate de novo and canonical biogenesis. Finally, we investigated which other factors modulate de novo centriole assembly and reveal that proteins of the pericentriolar matrix (PCM) promote biogenesis, likely by locally concentrating critical components.

Download Full-text

NEK7 is required for G1 progression and procentriole formation

Molecular Biology of the Cell ◽

10.1091/mbc.e16-09-0643 ◽

2017 ◽

Vol 28 (15) ◽

pp. 2123-2134 ◽

Cited By ~ 12

Author(s):

Akshari Gupta ◽

Yuki Tsuchiya ◽

Midori Ohta ◽

Gen Shiratsuchi ◽

Daiju Kitagawa

Keyword(s):

Cell Cycle ◽

Ubiquitin Ligase ◽

Cell Cycle Progression ◽

Primary Cilia ◽

S Phase ◽

Cycle Progression ◽

Restriction Point ◽

Centriole Duplication ◽

Abnormal Accumulation ◽

Phase Entry

The decision to commit to the cell cycle is made during G1 through the concerted action of various cyclin–CDK complexes. Not only DNA replication, but also centriole duplication is initiated as cells enter the S-phase. The NIMA-related kinase NEK7 is one of many factors required for proper centriole duplication, as well as for timely cell cycle progression. However, its specific roles in these events are poorly understood. In this study, we find that depletion of NEK7 inhibits progression through the G1 phase in human U2OS cells via down-regulation of various cyclins and CDKs and also inhibits the earliest stages of procentriole formation. Depletion of NEK7 also induces formation of primary cilia in human RPE1 cells, suggesting that NEK7 acts at least before the restriction point during G1. G1-arrested cells in the absence of NEK7 exhibit abnormal accumulation of the APC/C cofactor Cdh1 at the vicinity of centrioles. Furthermore, the ubiquitin ligase APC/CCdh1continuously degrades the centriolar protein STIL in these cells, thus inhibiting centriole assembly. Collectively our results demonstrate that NEK7 is involved in the timely regulation of G1 progression, S-phase entry, and procentriole formation.

Download Full-text

Loss of Primary Cilia in Melanoma Cells is Likely Independent of Proliferation and Cell Cycle Progression

Journal of Investigative Dermatology ◽

10.1038/jid.2015.22 ◽

2015 ◽

Vol 135 (5) ◽

pp. 1456-1458 ◽

Cited By ~ 15

Author(s):

Elizabeth R. Snedecor ◽

Clifford C. Sung ◽

Alejandra Moncayo ◽

Brooke E. Rothstein ◽

Daniel C. Mockler ◽

...

Keyword(s):

Cell Cycle ◽

Cell Cycle Progression ◽

Primary Cilia ◽

Melanoma Cells ◽

Cycle Progression

Download Full-text

An Overexpression Screen in Drosophila for Genes That Restrict Growth or Cell-Cycle Progression in the Developing Eye

Genetics ◽

10.1093/genetics/162.1.229 ◽

2002 ◽

Vol 162 (1) ◽

pp. 229-243 ◽

Cited By ~ 11

Author(s):

Ai-Sun Kelly Tseng ◽

Iswar K Hariharan

Keyword(s):

Cell Cycle ◽

Cell Proliferation ◽

Imaginal Disc ◽

Cell Cycle Progression ◽

Eye Development ◽

Gal4 Driver ◽

Proliferating Cells ◽

Cycle Progression ◽

Eye Imaginal Disc ◽

Doubling Times

AbstractWe screened for genes that, when overexpressed in the proliferating cells of the eye imaginal disc, result in a reduction in the size of the adult eye. After crossing the collection of 2296 EP lines to the ey-GAL4 driver, we identified 46 lines, corresponding to insertions in 32 different loci, that elicited a small eye phenotype. These lines were classified further by testing for an effect in postmitotic cells using the sev-GAL4 driver, by testing for an effect in the wing using en-GAL4, and by testing for the ability of overexpression of cycE to rescue the small eye phenotype. EP lines identified in the screen encompass known regulators of eye development including hh and dpp, known genes that have not been studied previously with respect to eye development, as well as 19 novel ORFs. Lines with insertions near INCENP, elB, and CG11518 were characterized in more detail with respect to changes in growth, cell-cycle phasing, and doubling times that were elicited by overexpression. RNAi-induced phenotypes were also analyzed in SL2 cells. Thus overexpression screens can be combined with RNAi experiments to identify and characterize new regulators of growth and cell proliferation.

Download Full-text