Ubiquitin-proteasome system controls ciliogenesis at the initial step of axoneme extension

Kasahara, Kousuke; Kawakami, Yoshihiro; Kiyono, Tohru; Yonemura, Shigenobu; Kawamura, Yoshifumi; Era, Saho; Matsuzaki, Fumio; Goshima, Naoki; Inagaki, Masaki

doi:10.1038/ncomms6081

Cited by 122 publications

(174 citation statements)

References 51 publications

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“…UBR5 was identified in a whole-genome shRNA screen as a positive regulator of ciliogenesis (90), with UBR5 depletion causing decreased primary cilia formation. This screening hit was not further validated and the mechanism underlying this effect is yet to be determined, but the UPS is an established regulator of ciliogenesis (91).…”

Section: Emerging Functional Roles For Ubr5mentioning

confidence: 99%

Functional Roles of the E3 Ubiquitin Ligase UBR5 in Cancer

Shearer

Iconomou

Watts

et al. 2015

Molecular Cancer Research

113

141

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The Ubiquitin-Proteasome System (UPS) is an important regulator of cell signaling and proteostasis, which are essential to a variety of cellular processes. The UPS is disrupted in many diseases including cancer, and targeting the UPS for cancer therapy is gaining wide interest. E3 ubiquitin ligases occupy a key position in the hierarchical UPS enzymatic cascade, largely responsible for determining substrate specificity and ubiquitin (Ub) chain topology. The E3 ligase UBR5 (aka EDD1) is emerging as a key regulator of the UPS in cancer and development. UBR5 expression is deregulated in many cancer types and UBR5 is frequently mutated in mantle cell lymphoma. UBR5 is highly conserved in metazoans, has unique structural features, and has been implicated in regulation of DNA damage response, metabolism, transcription, and apoptosis. Hence, UBR5 is a key regulator of cell signaling relevant to broad areas of cancer biology. However, the mechanism by which UBR5 may contribute to tumor initiation and progression remains poorly defined. This review synthesizes emerging insights from genetics, biochemistry, and cell biology to inform our understanding of UBR5 in cancer. These molecular insights indicate a role for UBR5 in integrating/coordinating various cellular signaling pathways. Finally, we discuss outstanding questions in UBR5 biology and highlight the need to systematically characterize substrates, and address limitations in current animal models, to better define the role of UBR5 in cancer.

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Section: Emerging Functional Roles For Ubr5mentioning

confidence: 99%

Functional Roles of the E3 Ubiquitin Ligase UBR5 in Cancer

Shearer

Iconomou

Watts

et al. 2015

Molecular Cancer Research

113

141

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“…Despite its name, KCTD17 has been shown to act as a substrate-adaptor for Cul3-RING ubiquitin ligases (CRL3s) to regulate proteosomal degradation of specific targets. 22, 36, 37 We observed that hepatic Kctd17 expression is robustly increased in HFD-fed and db/db mice (Figure 4A). Next, we confirmed our LC-MS/MS results by immunoprecipitation of endogenous KCTD17 in livers from HFD-fed mice with anti-PHLPP2 antibody, and vice versa (Figure 4B).…”

Section: Resultsmentioning

confidence: 89%

“…To understand the endogenous regulatory machinery that mediates PHLPP2 degradation in obese liver, we performed sequential LC/MS-MS-based screens, which identified novel glucagon/PKA-mediated PHLPP2 phosphorylation at Ser1119 and Ser1210, which augments PHLPP2 interaction with KCTD17, an adaptor protein for Cul3-RING ubiquitin ligases. 22 Interestingly, KCTD17 expression is increased in livers from obese mice and patients with NASH, linking glucagon-induced PHLPP2 phosphorylation with degradation. Thus, knockdown of KCTD17 in obese mice increases endogenous PHLPP2 and ameliorates obesity-induced fatty liver, revealing a novel strategy in the treatment of obesity-mediated NAFLD/NASH.…”

Section: Introductionmentioning

confidence: 99%

Degradation of PHLPP2 by KCTD17, via a Glucagon-Dependent Pathway, Promotes Hepatic Steatosis

et al. 2017

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BACKGROUND & AIMS Obesity-induced non-alcoholic fatty liver disease (NAFLD) develops, in part, via excess insulin-stimulated hepatic de novo lipogenesis, which increases, paradoxically, in patients with obesity-induced insulin resistance. Pleckstrin homology domain leucine-rich repeat protein phosphatase 2 (PHLPP2) terminates insulin signaling by dephosphorylating Akt; levels of PHLPP2 are reduced in livers from obese mice. We investigated whether loss of hepatic PHLPP2 is sufficient to induce fatty liver in mice, mechanisms of PHLPP2 degradation in fatty liver, and expression of genes that regulate PHLPP2 in livers of patients with NAFLD. METHODS C57BL/6J mice (controls), obese db/db mice and mice with liver-specific deletion of PHLPP2 (L-PHLPP2) fed either normal chow or high-fat diet (HFD) were analyzed for metabolic phenotypes including glucose tolerance and hepatic steatosis. PHLPP2-deficient primary hepatocytes or CRISPR/Cas9-mediated PHLPP2-knockout hepatoma cells were analyzed for insulin signaling and gene expression. We performed mass spectrometry analyses of livers tissues from C57BL/6J mice transduced with Ad-HA-FLAG-PHLPP2 to identify post-translational modifications to PHLPP2 and proteins that interact with PHLPP2. We measured levels of mRNAs by quantitative reverse transcription PCR in liver biopsies from patients with varying degrees of hepatic steatosis. RESULTS PHLPP2-knockout hepatoma cells and hepatocytes from L-PHLPP2 mice showed normal initiation of insulin signaling, but prolonged insulin action. Chow-fed L-PHLPP2 mice had normal glucose tolerance but hepatic steatosis. In HFD-fed C57BL/6J or db/db obese mice, endogenous PHLPP2 was degraded by glucagon and PKA-dependent phosphorylation of PHLPP2 (at Ser1119 and Ser1210), which led to PHLPP2 binding to potassium channel tetramerization domain containing 17 (KCTD17), a substrate-adaptor for Cul3-RING ubiquitin ligases. Levels of KCTD17 mRNA were increased in livers of HFD-fed C57BL/6J or db/db obese mice and in liver biopsies patients with NAFLD, compared with liver tissues from healthy control mice or patients without steatosis. Knockdown of KCTD17 with small hairpin RNA in primary hepatocytes increased PHLPP2 protein but not Phlpp2 mRNA, indicating that KCTD17 mediates PHLPP2 degradation. KCTD17 knockdown in obese mice prevented PHLPP2 degradation and decreased expression of lipogenic genes. CONCLUSIONS In mouse models of obesity, we found that PHLPP2 degradation induced lipogenesis without affecting gluconeogenesis. KCTD17, which is upregulated in liver tissues of obese mice and patients with NAFLD, binds to phosphorylated PHLPP2 to target it for ubiquitin-mediated degradation; this increases expression of genes that regulate lipogenesis to promote hepatic steatosis. Inhibitors of this pathway might be developed for treatment of patients with NAFLD.

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“…Given the results, we focused on the mechanisms by which cell cycle cues control ciliogenesis in the current study. Recent reports noted that proteolysis of ciliary proteins including Trichoplein contributes cell cycle exit and cilia formation (Tang et al 2013;Kasahara et al 2014). Therefore, it is conceivable that the defective formation of the cilia caused by the depletion of mRNA splicing regulators is due to deregulation of the cell cycle exit.…”

Section: Introductionmentioning

confidence: 99%

Mitotic control by mRNA splicing regulators ensures primary cilia formation

Kim

Lee

2016

Animal Cells and Systems

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The biogenesis of the primary cilium is coordinated with cell cycle exit/re-entry in most types of cells. After serum starvation, the cilia-generating cells enter quiescence and produce the primary cilium; upon re-addition of serum, they re-enter the cell cycle and resorb the cilium. We previously identified novel mechanisms to link cell cycle progression and ciliogenesis by highcontent genome-wide RNAi cell-based screening. In the present study, we pay attention to reveal the impact of mRNA splicing on cilia assembly after mitosis of cell cycle. We demonstrate that splicing regulators such as SON and XAB2 play an important role in mitosis exit, and thus affect ciliogenesis in G1/G0 phases. Knockdown of the splicing regulators in hTERT-RPE1 cells caused abnormal G2/M arrest under both serum addition and serum starvation, indicating defects in mitosis exit. Moreover, the knockdown cells failed to assemble the cilia under serum starvation and an inhibition of mRNA splicing using SSA, a spliceosome inhibitor, also revealed ciliogenesis defect. Finally, we show that the SSA-treated zebrafish display abnormal vascular development as a ciliary defect. These findings suggest the pivotal role of mRNA splicing regulators in cilia assembly and underscore the importance of mitotic regulation in ciliogenesis.ARTICLE HISTORY

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Ubiquitin-proteasome system controls ciliogenesis at the initial step of axoneme extension

Cited by 122 publications

References 51 publications

Functional Roles of the E3 Ubiquitin Ligase UBR5 in Cancer

Functional Roles of the E3 Ubiquitin Ligase UBR5 in Cancer

Degradation of PHLPP2 by KCTD17, via a Glucagon-Dependent Pathway, Promotes Hepatic Steatosis

Mitotic control by mRNA splicing regulators ensures primary cilia formation

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