Ubiquitin E3 Ligase Itch Negatively Regulates Osteoclast Formation by Promoting Deubiquitination of Tumor Necrosis Factor (TNF) Receptor-associated Factor 6

Zhang, Hengwei; Wu, Chengwu; Matesic, Lydia E.; Li, Xing; Wang, Zhiyu; Xing, Lianping

doi:10.1074/jbc.m112.442459

Cited by 30 publications

(55 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Itch transcription is directly regulated by RANKL during osteoclast differentiation via NF-κB. 14 In the current study, we demonstrated that NF-κB members also promote Itch expression in osteoblasts, indicating that NF-κB activation may affect protein ubiquitination and degradation in osteoblast lineage cells via Itch and perhaps other WW domain-containing E3 ligases.…”

Section: Discussionsupporting

confidence: 53%

“…14 Thus, we examined expression of NF-κB members at mRNA level extracted from callus tissues at different time points post-fracture. The expression levels of all NF-κB members, including p50, RelA, p52 and RelB, were significantly increased, starting at 7 days after fracture and reaching maximum expression at 14 days (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…14 We wanted to know if this also occurs in osteoblasts. We have also previously identified two putative NF-κB binding sites at the -3473/-3465 and -3183/-3175 location of the murine Itch promoter (National Centre for Biotechnology Information RefSeq accession number NC_000068) using TFSEARCH software (version 1.3, National Institute of Advanced Industrial Science and Technology, Tokyo).…”

Section: Resultsmentioning

confidence: 99%

“…We have also previously identified two putative NF-κB binding sites at the -3473/-3465 and -3183/-3175 location of the murine Itch promoter (National Centre for Biotechnology Information RefSeq accession number NC_000068) using TFSEARCH software (version 1.3, National Institute of Advanced Industrial Science and Technology, Tokyo). 14 We overexpressed RelA and RelB in C2C12 cells and found that both of them could increase Itch expression (Fig. 4a) and then performed ChIP assays using an anti- NF-κB RelA or RelB antibody to pull down protein-chromatin complexes and two primer sets around these NF-κB binding sites.…”

Section: Resultsmentioning

confidence: 99%

See 3 more Smart Citations

Ubiquitin E3 ligase Itch negatively regulates osteoblast function by promoting proteasome degradation of osteogenic proteins

Liu

Zhang

et al. 2017

Bone & Joint Research

View full text Add to dashboard Cite

ObjectivesUbiquitin E3 ligase-mediated protein degradation regulates osteoblast function. Itch, an E3 ligase, affects numerous cell functions by regulating ubiquitination and proteasomal degradation of related proteins. However, the Itch-related cellular and molecular mechanisms by which osteoblast differentiation and function are elevated during bone fracture repair are as yet unknown.MethodsWe examined the expression levels of E3 ligases and NF-κB members in callus samples during bone fracture repair by quantitative polymerase chain reaction (qPCR) and the total amount of ubiquitinated proteins by Western blot analysis in wild-type (WT) mice. The expression levels of osteoblast-associated genes in fracture callus from Itch knockout (KO) mice and their WT littermates were examined by qPCR. The effect of NF-κB on Itch expression in C2C12 osteoblast cells was determined by a chromatin immunoprecipitation (ChIP) assay.ResultsThe expression levels of WW Domain Containing E3 Ubiquitin Protein Ligase 1 (Wwp1), SMAD Specific E3 Ubiquitin Protein Ligase 1 (Smurf1), SMAD Specific E3 Ubiquitin Protein Ligase 2 (Smurf2) and Itch were all significantly increased in the fracture callus of WT mice, which was associated with elevated expression of NF-κB members and total ubiquitinated proteins. Callus tissue isolated from Itch KO mice expressed higher levels of osteoblast-associated genes, including Runx2, a positive regulator of osteoblast differentiation, but osteoclast-associated genes were not increased. Both NF-κB RelA and RelB proteins were found to bind to the NF-κB binding site in the mouse Itch promoter.ConclusionsOur findings indicate that Itch depletion may have a strong positive effect on osteoblast differentiation in fracture callus. Thus, ubiquitin E3 ligase Itch could be a potential target for enhancing bone fracture healing.Cite this article: J. Liu, X. Li, H. Zhang, R. Gu, Z. Wang, Z. Gao, L. Xing. Ubiquitin E3 ligase Itch negatively regulates osteoblast function by promoting proteasome degradation of osteogenic proteins. Bone Joint Res 2017;6:154–161. DOI: 10.1302/2046-3758.63.BJR-2016-0237.R1.

show abstract

Section: Discussionsupporting

confidence: 53%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

See 2 more Smart Citations

Ubiquitin E3 ligase Itch negatively regulates osteoblast function by promoting proteasome degradation of osteogenic proteins

Liu

Zhang

et al. 2017

Bone & Joint Research

View full text Add to dashboard Cite

show abstract

“…9 Itch is also required for negative regulation of TNF-and lipopolysaccharide (LPS)-mediated TNF receptor-associated factor 6 (TRAF6) ubiquitylation induced RING finger protein 11 10 and represent a negative regulator of osteoclastogenesis by promoting de-ubiquitylation of TRAF6. 11 Moreover, like other HECTs, Itch is also deregulated in cancer development and represents a potential target for anticancer treatment. 3,12 Since Itch is able to regulate chemosensitivity, 13 we attempted to develop specific inhibitors.…”

Section: Introductionmentioning

confidence: 99%

p63 threonine phosphorylation signals the interaction with the WW domain of the E3 ligase Itch

et al. 2014

View full text Add to dashboard Cite

Both in epithelial development as well as in epithelial cancers, the p53 family member p63 plays a crucial role acting as a master transcriptional regulator. P63 steady state protein levels are regulated by the E3 ubiquitin ligase Itch, via a physical interaction between the PPxY consensus sequence (PY motif) of p63 and one of the 4 WW domains of Itch; this substrate recognition process leads to protein-ubiquitylation and p63 proteasomal degradation. The interaction of the WW domains, a highly compact protein-protein binding module, with the short proline-rich sequences is therefore a crucial regulatory event that may offer innovative potential therapeutic opportunity. Previous molecular studies on the Itch-p63 recognition have been performed in vitro using the Itch-WW2 domain and the peptide interacting fragment of p63 (pep63), which includes the PY motif. Itch-WW2-pep63 interaction is also stabilized in vitro by the conformational constriction of the S-S cyclization in the p63 peptide. The PY motif of p63, as also for other proteins, is characterized by the nearby presence of a (T/S)P motif, which is a potential recognition site of the WW domain of the IV group present in the prolyl-isomerase Pin1. In this study, we demonstrate, by in silico and spectroscopical studies using both the linear pep63 and its cyclic form, that the threonine phosphorylation of the (T/S)PPPxY motif may represent a crucial regulatory event of the Itch-mediated p63 ubiquitylation, increasing the Itch-WW domains-p63 recognition event and stabilizing in vivo the Itch-WW-p63 complex. Moreover, our studies confirm that the subsequently trans/cis proline isomerization of (T/S)P motif by the Pin1 prolyl-isomerase, could modulate the E3-ligase interaction, and that the (T/S) p P trans PPxY motif represent the best conformer for the ItchWW-(T/S)PPPxY motif recognition.

show abstract

Regulation of RANKL‐induced osteoclastogenesis by RING finger protein RNF114

Lin

Qin

Leaman

et al. 2017

Journal Orthopaedic Research

View full text Add to dashboard Cite

Normal bone remodeling is a continuous process orchestrated by bone-resorbing osteoclasts and bone-forming osteoblasts, which an imbalance in bone remodeling results in metabolic bone diseases. RANKL, a member of the TNF cytokine family, functions as a key stimulator for osteoclast differentiation and maturation. Here, we report that RNF114, previously identified as a psoriasis susceptibility gene, plays a regulatory role in the RANKL/RANK/TRAF6 signaling pathway that mediates osteoclastogenesis. Our results demonstrated that RNF114 expression was significantly down-regulated in mouse osteoclast precursor cells undergoing RANKL-induced osteoclast differentiation. RNF114 knockout did not affect development or viability of the subpopulation of bone marrow macrophages capable of differentiating into osteoclasts in culture. However, in the presence of RANKL, RNF114 knockout bone marrow macrophages exhibited enhanced cell proliferation and augmented osteoclast differentiation, as shown by an increased expression of mature osteoclast markers, increased osteoclastic TRAP activity and bone resorption. Conversely, ectopic expression of RNF114 inhibited CTSK expression, TRAP activity, and bone resorption in RANKL-treated pre-osteoclasts. RNF114 also suppressed RANKL-activated NFATc1 expression and NFAT-regulated promoter activity. RNF114 suppressed TRAF6-, but not TAK1/TAB2-mediated NF-κB activation downstream of RANKL/RANK. In particular, TRAF6 protein levels were down-regulated by RNF114, possibly via K48-mediated proteasome-dependent degradation. These data suggested that RNF114's inhibitory effect on RANKL-stimulated osteoclastogenesis was mediated by blocking RANK/TRAF6/NF-κB signal transduction. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:159-166, 2018.

show abstract

Ubiquitin E3 Ligase Itch Negatively Regulates Osteoclast Formation by Promoting Deubiquitination of Tumor Necrosis Factor (TNF) Receptor-associated Factor 6

Abstract: Background: TRAF6 activity is crucial for osteoclastogenesis, which is decreased by deubiquitination. The role of Itch in this process is studied. Results: ItchϪ/Ϫ osteoclast precursors formed more osteoclasts, had prolonged RANKL-induced NF-B activation, and had

Cited by 30 publications

References 41 publications

Ubiquitin E3 ligase Itch negatively regulates osteoblast function by promoting proteasome degradation of osteogenic proteins

Ubiquitin E3 ligase Itch negatively regulates osteoblast function by promoting proteasome degradation of osteogenic proteins

p63 threonine phosphorylation signals the interaction with the WW domain of the E3 ligase Itch

Regulation of RANKL‐induced osteoclastogenesis by RING finger protein RNF114

Contact Info

Product

Resources

About