Ligand-dependent Degradation of Smad3 by a Ubiquitin Ligase Complex of ROC1 and Associated Proteins

Fukuchi, Minoru; Imamura, Takeshi; Chiba, Tomoki; Ebisawa, Takanori; Kawabata, Masahiro; Tanaka, Keiji; Miyazono, Kohei

doi:10.1091/mbc.12.5.1431

Cited by 201 publications

(157 citation statements)

References 50 publications

(71 reference statements)

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“…Moreover, although NEDD4-2 binds to both Smad2 and Smad3, which are structurally similar to each other (92 % identity), NEDD4-2 induces ubiquitin-dependent degradation of Smad2, but does not efficiently induce that of Smad3. We have reported previously that ROC1-SCF Fbw1a is the ubiquitin E3 ligase for Smad3 in TGF-β signalling [11]. Thus different types of E3 ubiquitin ligases, e.g.…”

Section: Discussionmentioning

confidence: 99%

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NEDD4-2 (neural precursor cell expressed, developmentally down-regulated 4-2) negatively regulates TGF-β (transforming growth factor-β) signalling by inducing ubiquitin-mediated degradation of Smad2 and TGF-β type I receptor

Kuratomi

Komuro

Goto

et al. 2005

Biochemical Journal

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197

143

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Inhibitory Smad, Smad7, is a potent inhibitor of TGF-β (transforming growth factor-β) superfamily signalling. By binding to activated type I receptors, it prevents the activation of R-Smads (receptor-regulated Smads). To identify new components of the Smad pathway, we performed yeast two-hybrid screening using Smad7 as bait, and identified NEDD4-2 (neural precursor cell expressed, developmentally down-regulated 4-2) as a direct binding partner of Smad7. NEDD4-2 is structurally similar to Smurfs (Smad ubiquitin regulatory factors) 1 and 2, which were identified previously as E3 ubiquitin ligases for R-Smads and TGF-β superfamily receptors. NEDD4-2 functions like Smurfs 1 and 2 in that it associates with TGF-β type I receptor via Smad7, and induces its ubiquitin-dependent degradation. Moreover, NEDD4-2 bound to TGF-β-specific R-Smads, Smads 2 and 3, in a ligand-dependent manner, and induced degradation of Smad2, but not Smad3. However, in contrast with Smurf2, NEDD4-2 failed to induce ubiquitination of SnoN (Ski-related novel protein N), although NEDD4-2 bound to SnoN via Smad2 more strongly than Smurf2. We showed further that overexpressed NEDD4-2 prevents transcriptional activity induced by TGF-β and BMP, whereas silencing of the NEDD4-2 gene by siRNA (small interfering RNA) resulted in enhancement of the responsiveness to TGF-β superfamily cytokines. These data suggest that NEDD4-2 is a member of the Smurf-like C2-WW-HECT (WW is Trp-Trp and HECT is homologous to the E6-accessory protein) type E3 ubiquitin ligases, which negatively regulate TGF-β superfamily signalling through similar, but not identical, mechanisms to those used by Smurfs.

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Section: Discussionmentioning

confidence: 99%

“…A RING-type E3 ubiquitin ligase ROC1-SCF (regulator of Cullins 1-Skp1/Cullin1/Fbox protein) complex was shown to degrade activated Smad3 and Smad4 [11,12]. Smurf (Smad ubiquitin regulatory factor) 1 was originally identified as a HECT-type E3 ubiquitin ligase for BMPspecific R-Smads, Smads 1 and 5 [13].…”

Section: Introductionmentioning

confidence: 99%

NEDD4-2 (neural precursor cell expressed, developmentally down-regulated 4-2) negatively regulates TGF-β (transforming growth factor-β) signalling by inducing ubiquitin-mediated degradation of Smad2 and TGF-β type I receptor

Kuratomi

Komuro

Goto

et al. 2005

Biochemical Journal

Self Cite

197

143

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“…Although p53 lacks a consensus DS(p)GXXS(p) phosphodependent binding motif for ␤-TrCP, binding of p53 to ␤-TrCP1 requires phosphorylation. ␤-TrCP has been identified as the E3 ligase for ubiquitination of Wee1, p100, and Smad3, all of which lack the consensus binding motif for ␤-TrCP (29)(30)(31). Therefore, it is reasonable to propose that IKK2 phosphorylation at serines 362 and 366 targets p53 for ubiquitination by ␤-TrCP1.…”

Section: Discussionmentioning

confidence: 99%

Phosphorylation of p53 by IκB kinase 2 promotes its degradation by β-TrCP

Xia

Padre

Mendoza

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

141

122

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Functional inactivation of p53 and constitutive activation of the NF-B pathway has been associated with several human cancers. In this study, we show that I B kinase 2 (IKK2/IKK␤), which is critical for NF-B activation, also phosphorylates p53. Phosphorylation of p53 at serines 362 and 366 by IKK2 leads to its recruitment to and ubiquitination by ␤-TrCP1. Degradation of ubiquitinated p53 is independent of Mdm2, because it occurs in both wild-type and Mdm2 ؊/؊ cells. SiRNA-mediated reduction in the levels of ␤-TrCP1 and other members of the SCF ␤؊TrCP1 E3 ubiquitin ligase complex or overexpression of a dominant negative form of ␤-TrCP1 enhances p53 stability. Substitutions at Ser-362 and 366 of p53 by alanines (p53 AA) result in reduced phosphorylation of p53 by IKK2, decreased association with ␤-TrCP1, and thus increased stability of p53 and expression of p53 target genes such as p21, altering the G1 phase of the cell cycle. Our results identify IKK2 and ␤-TrCP1 as novel regulators of the p53 pathway and suggest that blocking of IKK2 and ␤-TrCP1 could be a means of regulating p53 stability and thereby modulating its biological activity.␤-TrCP1 ͉ DNA damage ͉ IKK2 ͉ p53 stability N F-B is a family of transcription factors crucial for several biological processes involved in innate and adaptive immunity, inflammation, cell survival, and cancer (1). More than 150 different types of stimuli, including proinflammatory cytokines, bacterial/ viral infection, and stress, activate NF-B-dependent transcription of target genes. Activation of the I B kinase (IKK) complex is a prerequisite for NF-B activation. Once activated, IKK phosphorylates serines 32 and 36 of I B proteins (2), followed by ubiquitination by ␤-TrCP1, an E3 ubiquitin ligase, which targets them for proteosomal degradation (3). IKK2 has been shown to phosphorylate other cellular and viral proteins, such as ␤-catenin, I B␤, I B , p65, HIV-VPU, and SRC-3; this phosphorylation regulates the steady-state levels and transcriptional activity of these substrates (4-7). Approximately 30% of cellular proteins contain covalently bound phosphate, even though only 500-600 protein kinases likely are encoded by the human genome (8). Therefore, it is conceivable that, like many other kinases, IKK2 also has additional substrates, and that each protein phosphorylated by IKK2 in turn can be a substrate of other kinases.The p53 tumor suppressor plays a crucial role in the development of many cancers and is frequently mutated or deleted in human cancers (9, 10). In the absence of stress, endogenous p53 levels are very low, because of the constant recruitment of p53 to Mdm2, an E3 ubiquitin ligase that inhibits its transcriptional activity and targets it for proteosomal degradation (11,12). In response to DNA damage and other cellular stresses, the p53 protein levels are up-regulated and its activities are induced. The inhibition of Mdm2 binding to p53 and increases in p53 transcriptional activity involve the phosphorylation and stabilization of p53.Human p53 has been reported ...

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“…Smad3 is not directly affected by the Smurfs even though it contains the PY motif and binds Smurfs. Instead, the SCF/Roc1 E3 Ub ligase complex controls degradation of activated phospho-Smad3 [53]. SCF/ Roc1 is suggested to terminate Smad3 nuclear signaling by ubiquitinating Smad3 and favoring its export to the cytoplasm for proteasomal degradation.…”

Section: Regulation Of R-smad Stabilitymentioning

confidence: 99%

Regulating the stability of TGFβ receptors and Smads

et al. 2008

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Abbreviations: AIP4 (atrophin 1-interacting protein 4); BMP (bone morphogenetic protein); CHIP (carboxyl terminus of Hsc70-interacting protein); GSK3b (glycogen synthase kinase 3-b); HECT (homologous to E6-AP carboxyl terminus); MAPK (mitogen-activated protein kinase); NEDD4-2 (neural precursor cell expressed, developmentally downregulated 4-2); PIAS (protein inhibitor of activated Stat); Smurf (Smad ubiquitylation regulatory factor); STRAP (serine-threonine kinase receptorassociated protein); SUMO (small ubiquitin-like modifier); TbRI/TbRII (TGFb receptor type I and II); TGFb (transforming growth factor b); WWP1 (WW domain-containing protein 1); Ub (ubiquitin) npg Transforming growth factor β (TGFβ) controls cellular behavior in embryonic and adult tissues. TGFβ binding to serine/threonine kinase receptors on the plasma membrane activates Smad molecules and additional signaling proteins that together regulate gene expression. In this review, mechanisms and models that aim at explaining the coordination between several components of the signaling network downstream of TGFβ are presented. We discuss how the activity and duration of TGFβ receptor/Smad signaling can be regulated by post-translational modifications that affect the stability of key proteins in the pathway. We highlight links between these mechanisms and human diseases, such as tissue fibrosis and cancer. Regulating the stability of TGFβ receptors and Smads

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Ligand-dependent Degradation of Smad3 by a Ubiquitin Ligase Complex of ROC1 and Associated Proteins

Cited by 201 publications

References 50 publications

NEDD4-2 (neural precursor cell expressed, developmentally down-regulated 4-2) negatively regulates TGF-β (transforming growth factor-β) signalling by inducing ubiquitin-mediated degradation of Smad2 and TGF-β type I receptor

NEDD4-2 (neural precursor cell expressed, developmentally down-regulated 4-2) negatively regulates TGF-β (transforming growth factor-β) signalling by inducing ubiquitin-mediated degradation of Smad2 and TGF-β type I receptor

Phosphorylation of p53 by IκB kinase 2 promotes its degradation by β-TrCP

Regulating the stability of TGFβ receptors and Smads

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