scholarly journals C3G (RapGEF1) localizes to the mother centriole and regulates centriole division and primary cilia dynamics

2019 ◽  
Author(s):  
Sanjeev Chavan Nayak ◽  
Vegesna Radha
Keyword(s):  
Embryonic Development ◽  
Primary Cilia ◽  
Negative Regulator ◽  
Ciliated Cells ◽  
Wild Type ◽  
Centrosomal Protein ◽  
Over Expression ◽  
Cilia Formation ◽  
Mother Centriole ◽  
Summary Statement

AbstractC3G (RapGEF1), a negative regulator of β-catenin, plays a role in cell differentiation and is essential for early embryonic development in mice. In this study, we identify C3G as a centrosomal protein that regulates centriole division and primary cilia dynamics. C3G is present at the centrosome in interphase as well as mitotic cells, but is absent at the centrioles in differentiated myotubes. It interacts with, and co-localizes with cenexin in the mother centriole. Stable clone of cells depleted of C3G by CRISPR/Cas9 showed reduction in cenexin protein, and presence of supernumerary centrioles. Over-expression of C3G resulted in inhibition of centrosome division in normal and hydroxyurea treated cells. Proportion of ciliated cells is higher, and cilia length longer in C3G knockout cells. C3G inhibits cilia formation and length dependent on its catalytic activity. Unlike wild type cells, C3G depleted cells inefficiently retracted their cilia upon stimulation to reenter the cell cycle, and proliferated slowly, arresting in G1. We conclude that C3G inhibits centriole division and maintains ciliary homeostasis, properties that may be important for its role in embryonic development.Summary statementWe identify C3G as a centrosomal protein and regulator of centriole number, primary cilia length and resorption. These properties are important for its role in embryogenesis, and suggest that mutations in C3G could cause ciliopathies.

scholarly journals CEP97 phosphorylation by Dyrk1a is critical for centriole separation during multiciliogenesis

2021 ◽  
Vol 221 (1) ◽  
Author(s):  
Moonsup Lee ◽  
Kunio Nagashima ◽  
Jaeho Yoon ◽  
Jian Sun ◽  
Ziqiu Wang ◽  
...  
Keyword(s):  
Embryonic Development ◽  
Tyrosine Phosphorylation ◽  
Primary Cilia ◽  
Structural Characteristics ◽  
Xenopus Embryo ◽  
Negative Regulator ◽  
Dual Specificity ◽  
Multiciliated Cells ◽  
Cilia Formation

Proper cilia formation in multiciliated cells (MCCs) is necessary for appropriate embryonic development and homeostasis. Multicilia share many structural characteristics with monocilia and primary cilia, but there are still significant gaps in our understanding of the regulation of multiciliogenesis. Using the Xenopus embryo, we show that CEP97, which is known as a negative regulator of primary cilia formation, interacts with dual specificity tyrosine phosphorylation regulated kinase 1A (Dyrk1a) to modulate multiciliogenesis. We show that Dyrk1a phosphorylates CEP97, which in turn promotes the recruitment of Polo-like kinase 1 (Plk1), which is a critical regulator of MCC maturation that functions to enhance centriole disengagement in cooperation with the enzyme Separase. Knockdown of either CEP97 or Dyrk1a disrupts cilia formation and centriole disengagement in MCCs, but this defect is rescued by overexpression of Separase. Thus, our study reveals that Dyrk1a and CEP97 coordinate with Plk1 to promote Separase function to properly form multicilia in vertebrate MCCs.

scholarly journals CEP19 cooperates with FOP and CEP350 to drive early steps in the ciliogenesis programme

Open Biology ◽  
2017 ◽  
Vol 7 (6) ◽  
pp. 170114 ◽  
Author(s):  
Bahareh A. Mojarad ◽  
Gagan D. Gupta ◽  
Monica Hasegan ◽  
Oumou Goudiam ◽  
Renata Basto ◽  
...  
Keyword(s):  
Primary Cilia ◽  
Three Dimensional ◽  
Structured Illumination ◽  
Loss Of Function ◽  
Structured Illumination Microscopy ◽  
C Terminus ◽  
Interaction Partners ◽  
Cilia Formation ◽  
Mother Centriole ◽  
Section Electron

Primary cilia are microtubule-based sensory organelles necessary for efficient transduction of extracellular cues. To initiate cilia formation, ciliary vesicles (CVs) are transported to the vicinity of the centrosome where they dock to the distal end of the mother centriole and fuse to initiate cilium assembly. However, to this date, the early steps in cilia formation remain incompletely understood. Here, we demonstrate functional interplay between CEP19, FOP and CEP350 in ciliogenesis. Using three-dimensional structured-illumination microscopy (3D-SIM) imaging, we mapped the relative spatial distribution of these proteins at the distal end of the mother centriole and show that CEP350/FOP act upstream of CEP19 in their recruitment hierarchy. We demonstrate that CEP19 CRISPR KO cells are severely impaired in their ability to form cilia, analogous to the loss of function of CEP19 binding partners FOP and CEP350. Notably, in the absence of CEP19 microtubule anchoring at centromes is similar in manner to its interaction partners FOP and CEP350. Using GFP-tagged deletion constructs of CEP19, we show that the C-terminus of CEP19 is required for both its localization to centrioles and for its function in ciliogenesis. Critically, this region also mediates the interaction between CEP19 and FOP/CEP350. Interestingly, a morbid-obesity-associated R82* truncated mutant of CEP19 cannot ciliate nor interact with FOP and CEP350, indicative of a putative role for CEP19 in ciliopathies. Finally, analysis of CEP19 KO cells using thin-section electron microscopy revealed marked defects in the docking of CVs to the distal end of the mother centrioles. Together, these data demonstrate a role for the CEP19, FOP and CEP350 module in ciliogenesis and the possible effect of disrupting their functions in ciliopathies.

scholarly journals Ndel1 suppresses ciliogenesis in proliferating cells by regulating the trichoplein–Aurora A pathway

2016 ◽  
Vol 212 (4) ◽  
pp. 409-423 ◽  
Author(s):  
Hironori Inaba ◽  
Hidemasa Goto ◽  
Kousuke Kasahara ◽  
Kanako Kumamoto ◽  
Shigenobu Yonemura ◽  
...  
Keyword(s):  
Primary Cilia ◽  
Serum Starvation ◽  
Aurora A ◽  
Proliferating Cells ◽  
Quiescent Cells ◽  
Cell Cycle Reentry ◽  
Upstream Regulator ◽  
Cilia Formation ◽  
Mother Centriole ◽  
Component Protein

Primary cilia protrude from the surface of quiescent cells and disassemble at cell cycle reentry. We previously showed that ciliary reassembly is suppressed by trichoplein-mediated Aurora A activation pathway in growing cells. Here, we report that Ndel1, a well-known modulator of dynein activity, localizes at the subdistal appendage of the mother centriole, which nucleates a primary cilium. In the presence of serum, Ndel1 depletion reduces trichoplein at the mother centriole and induces unscheduled primary cilia formation, which is reverted by forced trichoplein expression or coknockdown of KCTD17 (an E3 ligase component protein for trichoplein). Serum starvation induced transient Ndel1 degradation, subsequent to the disappearance of trichoplein at the mother centriole. Forced expression of Ndel1 suppressed trichoplein degradation and axonemal microtubule extension during ciliogenesis, similar to trichoplein induction or KCTD17 knockdown. Most importantly, the proportion of ciliated and quiescent cells was increased in the kidney tubular epithelia of newborn Ndel1-hypomorphic mice. Thus, Ndel1 acts as a novel upstream regulator of the trichoplein–Aurora A pathway to inhibit primary cilia assembly.

scholarly journals The Talpid3 gene (KIAA0586) encodes a centrosomal protein that is essential for primary cilia formation

Development ◽  
2009 ◽  
Vol 136 (4) ◽  
pp. 655-664 ◽  
Author(s):  
Y. Yin ◽  
F. Bangs ◽  
I. R. Paton ◽  
A. Prescott ◽  
J. James ◽  
...  
Keyword(s):  
Primary Cilia ◽  
Centrosomal Protein ◽  
Cilia Formation

scholarly journals Genotoxic stress-activated DNA-PK-p53 cascade and autophagy cooperatively induce ciliogenesis to maintain the DNA damage response

2021 ◽  
Author(s):  
Ting-Yu Chen ◽  
Bu-Miin Huang ◽  
Tang K. Tang ◽  
Yu-Ying Chao ◽  
Xiao-Yi Xiao ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Damage Response ◽  
Primary Cilium ◽  
Primary Cilia ◽  
Genotoxic Stress ◽  
Dna Damage Signaling ◽  
Damage Response ◽  
Cilia Formation ◽  
Mother Centriole ◽  
Autophagy Inhibition

AbstractThe DNA-PK maintains cell survival when DNA damage occurs. In addition, aberrant activation of the DNA-PK induces centrosome amplification, suggesting additional roles for this kinase. Here, we showed that the DNA-PK-p53 cascade induced primary cilia formation (ciliogenesis), thus maintaining the DNA damage response under genotoxic stress. Treatment with genotoxic drugs (etoposide, neocarzinostatin, hydroxyurea, or cisplatin) led to ciliogenesis in human retina (RPE1), trophoblast (HTR8), lung (A459), and mouse Leydig progenitor (TM3) cell lines. Upon genotoxic stress, several DNA damage signaling were activated, but only the DNA-PK-p53 cascade contributed to ciliogenesis, as pharmacological inhibition or genetic depletion of this pathway decreased genotoxic stress-induced ciliogenesis. Interestingly, in addition to localizing to the nucleus, activated DNA-PK localized to the base of the primary cilium (mother centriole) and daughter centriole. Genotoxic stress also induced autophagy. Inhibition of autophagy initiation or lysosomal degradation or depletion of ATG7 decreased genotoxic stress-induced ciliogenesis. Besides, inhibition of ciliogenesis by depletion of IFT88 or CEP164 attenuated the genotoxic stress-induced DNA damage response. Thus, our study uncovered the interplay among genotoxic stress, the primary cilium, and the DNA damage response.

scholarly journals CEP78 functions downstream of CEP350 to control biogenesis of primary cilia by negatively regulating CP110 levels

2020 ◽  
Author(s):  
André Brás Gonçalves ◽  
Beinta Biskopstø Joensen ◽  
Sarah Kirstine Hasselbalch ◽  
Pernille Martens ◽  
Signe Krogh Ohlsen ◽  
...  
Keyword(s):  
Hearing Loss ◽  
Ubiquitin Ligase ◽  
Primary Cilia ◽  
E3 Ubiquitin Ligase ◽  
Centrosomal Protein ◽  
Ubiquitin Ligase Complex ◽  
Cilia Formation ◽  
Retinal Cone ◽  
Dependent Mechanism

AbstractCEP78 is a centrosomal protein implicated in ciliogenesis and ciliary length control, and mutations in the CEP78 gene cause retinal cone-rod dystrophy associated with hearing loss. However, the mechanism by which CEP78 affects cilia formation is unknown. Based on a recently-discovered disease-causing CEP78 p.L150S mutation, we identified the disease-relevant interactome of CEP78. We confirmed that CEP78 interacts with the EDD1-DYRK2-DDB1VPRBP E3 ubiquitin ligase complex, which is involved in CP110 ubiquitination and degradation, and identified a novel interaction between CEP78 and CEP350 that is weakened by the CEP78L150S mutation. We show that CEP350 promotes centrosomal recruitment and stability of CEP78, which in turn leads to centrosomal recruitment of EDD1. Consistently, cells lacking CEP78 display significantly increased cellular and centrosomal levels of CP110, and depletion of CP110 in CEP78-deficient cells restored ciliation frequency to normal. We propose that CEP78 functions downstream of CEP350 to promote ciliogenesis by negatively regulating CP110 levels via an EDD1-dependent mechanism.

scholarly journals NudCL2 is an autophagy receptor that mediates selective autophagic degradation of CP110 at mother centrioles to promote ciliogenesis

Cell Research ◽  
2021 ◽  
Author(s):  
Min Liu ◽  
Wen Zhang ◽  
Min Li ◽  
Jiaxing Feng ◽  
Wenjun Kuang ◽  
...  
Keyword(s):  
Primary Cilia ◽  
Coiled Coil ◽  
Serum Starvation ◽  
Vertebrate Development ◽  
Wild Type ◽  
Selective Degradation ◽  
Mother Centriole ◽  
Autophagic Degradation ◽  
Specific Degradation ◽  
Lir Motif

AbstractPrimary cilia extending from mother centrioles are essential for vertebrate development and homeostasis maintenance. Centriolar coiled-coil protein 110 (CP110) has been reported to suppress ciliogenesis initiation by capping the distal ends of mother centrioles. However, the mechanism underlying the specific degradation of mother centriole-capping CP110 to promote cilia initiation remains unknown. Here, we find that autophagy is crucial for CP110 degradation at mother centrioles after serum starvation in MEF cells. We further identify NudC-like protein 2 (NudCL2) as a novel selective autophagy receptor at mother centrioles, which contains an LC3-interacting region (LIR) motif mediating the association of CP110 and the autophagosome marker LC3. Knockout of NudCL2 induces defects in the removal of CP110 from mother centrioles and ciliogenesis, which are rescued by wild-type NudCL2 but not its LIR motif mutant. Knockdown of CP110 significantly attenuates ciliogenesis defects in NudCL2-deficient cells. In addition, NudCL2 morphants exhibit ciliation-related phenotypes in zebrafish, which are reversed by wild-type NudCL2, but not its LIR motif mutant. Importantly, CP110 depletion significantly reverses these ciliary phenotypes in NudCL2 morphants. Taken together, our data suggest that NudCL2 functions as an autophagy receptor mediating the selective degradation of mother centriole-capping CP110 to promote ciliogenesis, which is indispensable for embryo development in vertebrates.

scholarly journals CEP78 functions downstream of CEP350 to control biogenesis of primary cilia by negatively regulating CP110 levels

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
André B Goncalves ◽  
Sarah K Hasselbalch ◽  
Beinta B Joensen ◽  
Sebastian Patzke ◽  
Pernille Martens ◽  
...  
Keyword(s):  
Hearing Loss ◽  
Ubiquitin Ligase ◽  
Primary Cilia ◽  
E3 Ubiquitin Ligase ◽  
Centrosomal Protein ◽  
Ubiquitin Ligase Complex ◽  
Cilia Formation ◽  
Retinal Cone ◽  
Dependent Mechanism

CEP78 is a centrosomal protein implicated in ciliogenesis and ciliary length control, and mutations in the CEP78 gene cause retinal cone-rod dystrophy associated with hearing loss. However, the mechanism by which CEP78 affects cilia formation is unknown. Based on a recently discovered disease-causing CEP78 p.L150S mutation, we identified the disease-relevant interactome of CEP78. We confirmed that CEP78 interacts with the EDD1-DYRK2-DDB1VPRBP E3 ubiquitin ligase complex, which is involved in CP110 ubiquitination and degradation, and identified a novel interaction between CEP78 and CEP350 that is weakened by the CEP78L150S mutation. We show that CEP350 promotes centrosomal recruitment and stability of CEP78, which in turn leads to centrosomal recruitment of EDD1. Consistently, cells lacking CEP78 display significantly increased cellular and centrosomal levels of CP110, and depletion of CP110 in CEP78-deficient cells restored ciliation frequency to normal. We propose that CEP78 functions downstream of CEP350 to promote ciliogenesis by negatively regulating CP110 levels via an EDD1-dependent mechanism.

scholarly journals Trisomy 21 increases microtubules and disrupts centriolar satellite localization

2021 ◽  
Author(s):  
Bailey L McCurdy ◽  
Cayla E Jewett ◽  
Alexander J StemmWolf ◽  
Huy Nguyen Duc ◽  
Molishree Joshi ◽  
...  
Keyword(s):  
Down Syndrome ◽  
Intracellular Trafficking ◽  
Trisomy 21 ◽  
Primary Cilia ◽  
Chromosome 21 ◽  
Normal Density ◽  
Wild Type ◽  
Microtubule Network ◽  
Cilia Formation

Trisomy 21, the cause of Down syndrome, causes a 0.5-fold protein increase of the chromosome 21-resident gene Pericentrin (PCNT) and reduces primary cilia formation and signaling. Here we investigate the mechanisms by which PCNT imbalances disrupt cilia. Using isogenic RPE-1 cells with increased chromosome 21 dosage, we find PCNT protein accumulates around the centrosome as a pericentrosomal cluster of enlarged cytoplasmic puncta that localize along and at MT ends. Cytoplasmic PCNT puncta impact the intracellular MT trafficking network required for primary cilia, as the PCNT puncta sequester cargo peripheral to centrosomes in what we call pericentrosomal crowding. The centriolar satellite proteins, PCM1, CEP131 and CEP290, important for ciliogenesis, accumulate at sites of enlarged PCNT puncta in trisomy 21 cells. Reducing PCNT when chromosome 21 ploidy is elevated is sufficient to decrease PCNT puncta, reestablish a normal density of MTs around the centrosome, restore ciliogenesis to wild type levels and decrease pericentrosomal crowding. A transient reduction in MTs also decreases pericentrosomal crowding and partially rescues ciliogenesis in trisomy 21 cells, indicating that increased PCNT leads to defects in the microtubule network deleterious to normal centriolar satellite distribution. We propose that chromosome 21 aneuploidy disrupts MT-dependent intracellular trafficking required for primary cilia.

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