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Sharad Kumar - One of the best experts on this subject based on the ideXlab platform.
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Dietary sodium modulates nephropathy in Nedd4-2-deficient mice
Cell Death & Differentiation, 2020Co-Authors: Jantina A Manning, Sonia S. Shah, Tanya L. Henshall, Andrej Nikolic, John Finnie, Sharad KumarAbstract:Salt homeostasis is maintained by tight control of Na^+ filtration and reabsorption. In the distal part of the nephron the ubiquitin protein ligase Nedd4-2 regulates membrane abundance and thus activity of the epithelial Na^+ channel (ENaC), which is rate-limiting for Na^+ reabsorption. Nedd4-2 deficiency in mouse results in elevated ENaC and nephropathy, however the contribution of dietary salt to this has not been characterized. In this study we show that high dietary Na^+ exacerbated kidney injury in Nedd4-2 -deficient mice, significantly perturbing normal postnatal nephrogenesis and resulting in multifocal areas of renal dysplasia, increased markers of kidney injury and a decline in renal function. In control mice, high dietary Na^+ resulted in reduced levels of ENaC. However, Nedd4-2 -deficient kidneys maintained elevated ENaC even after high dietary Na^+, suggesting that the inability to efficiently downregulate ENaC is responsible for the salt-sensitivity of disease. Importantly, low dietary Na^+ significantly ameliorated nephropathy in Nedd4-2 -deficient mice. Our results demonstrate that due to dysregulation of ENaC, kidney injury in Nedd4-2 -deficient mice is sensitive to dietary Na^+, which may have implications in the management of disease in patients with kidney disease.
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nedd4 2 NEDD4L the ubiquitin ligase for multiple membrane proteins
Gene, 2015Co-Authors: Pranay Goel, Jantina A Manning, Sharad KumarAbstract:Abstract NEDD4-2 (also known as NEDD4L, neural precursor cell expressed developmentally down-regulated 4-like) is a ubiquitin protein ligase of the Nedd4 family which is known to bind and regulate a number of membrane proteins to aid in their internalization and turnover. Several of the NEDD4-2 substrates include ion channels, such as the epithelial and voltage-gated sodium channels. Given the critical function of NEDD4-2 in regulating membrane proteins, this ligase is essential for the maintenance of cellular homeostasis. In this article we review the biology and function of this important ubiquitin-protein ligase and discuss its pathophysiological significance.
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nedd4 2 NEDD4L controls intracellular na mediated activity of voltage gated sodium channels in primary cortical neurons
Biochemical Journal, 2014Co-Authors: Jenny Ekberg, Natasha A. Boase, Jantina A Manning, Sharad Kumar, Grigori Y Rychkov, Philip PoronnikAbstract:Nedd4-2, a HECT (homologous with E6-associated protein C-terminus)-type ubiquitin protein ligase, has been implicated in regulating several ion channels, including Navs (voltage-gated sodium channels). In Xenopus oocytes Nedd4-2 strongly inhibits the activity of multiple Navs. However, the conditions under which Nedd4-2 mediates native Nav regulation remain uncharacterized. Using Nedd4-2-deficient mice, we demonstrate in the present study that in foetal cortical neurons Nedd4-2 regulates Navs specifically in response to elevated intracellular Na(+), but does not affect steady-state Nav activity. In dorsal root ganglia neurons from the same mice, however, Nedd4-2 does not control Nav activities. The results of the present study provide the first physiological evidence for an essential function of Nedd4-2 in regulating Navs in the central nervous system.
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Nedd4-2 (NEDD4L) controls intracellular Na+-mediated activity of voltage-gated sodium channels in primary cortical neurons
Biochemical Journal, 2013Co-Authors: Jenny Ekberg, Natasha A. Boase, Jantina A Manning, Grigori Y Rychkov, Philip Poronnik, Sharad KumarAbstract:Nedd4-2, a HECT (homologous with E6-associated protein C-terminus)-type ubiquitin protein ligase, has been implicated in regulating several ion channels, including Navs (voltage-gated sodium channels). In Xenopus oocytes Nedd4-2 strongly inhibits the activity of multiple Navs. However, the conditions under which Nedd4-2 mediates native Nav regulation remain uncharacterized. Using Nedd4-2-deficient mice, we demonstrate in the present study that in foetal cortical neurons Nedd4-2 regulates Navs specifically in response to elevated intracellular Na(+), but does not affect steady-state Nav activity. In dorsal root ganglia neurons from the same mice, however, Nedd4-2 does not control Nav activities. The results of the present study provide the first physiological evidence for an essential function of Nedd4-2 in regulating Navs in the central nervous system.
Chia Hsiung Cheng - One of the best experts on this subject based on the ideXlab platform.
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igf 1 enhanced mir 513a 5p signaling desensitizes glioma cells to temozolomide by targeting the NEDD4L inhibited wnt β catenin pathway
PLOS ONE, 2019Co-Authors: Ku Chung Chen, Peng Hsu Chen, Kuo Hao Ho, Chwen Ming Shih, Chih Ming Chou, Chia Hsiung ChengAbstract:: Temozolomide (TMZ) is a first-line alkylating agent for glioblastoma multiforme (GBM). Clarifying the mechanisms inducing TMZ insensitivity may be helpful in improving its therapeutic effectiveness against GBM. Insulin-like growth factor (IGF)-1 signaling and micro (mi)RNAs are relevant in mediating GBM progression. However, their roles in desensitizing GBM cells to TMZ are still unclear. We aimed to identify IGF-1-mediated miRNA regulatory networks that elicit TMZ insensitivity for GBM. IGF-1 treatment attenuated TMZ cytotoxicity via WNT/β-catenin signaling, but did not influence glioma cell growth. By miRNA array analyses, 93 upregulated and 148 downregulated miRNAs were identified in IGF-1-treated glioma cells. miR-513a-5p from the miR-513a-2 gene locus was upregulated by IGF-1-mediated phosphoinositide 3-kinase (PI3K) signaling. Its elevated levels were also observed in gliomas versus normal cells, in array data of The Cancer Genome Atlas (TCGA), and the GSE61710, GSE37366, and GSE41032 datasets. In addition, lower levels of neural precursor cell-expressed developmentally downregulated 4-like (NEDD4L), an E3 ubiquitin protein ligase that inhibits WNT signaling, were found in gliomas by analyzing cells, arrays, and RNA sequencing data of TCGA glioma patients. Furthermore, a negative correlation was identified between miR-513a-5p and NEDD4L in glioma. NEDD4L was also validated as a direct target gene of miR-513a-5p, and it was reduced by IGF-1 treatment. Overexpression of NEDD4L inhibited glioma cell viability and reversed IGF-1-repressed TMZ cytotoxicity. In contrast, miR-513a-5p significantly affected NEDD4L-inhibited WNT signaling and reduced TMZ cytotoxicity. These findings demonstrate a distinct role of IGF-1 signaling through miR-513a-5p-inhibited NEDD4L networks in influencing GBM's drug sensitivity to TMZ.
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IGF-1-enhanced miR-513a-5p Signaling Desensitizes Glioma Cells to Temozolomide by Targeting the NEDD4L-inhibited Wnt/β-catenin Pathway
PLOS ONE, 2019Co-Authors: Ku Chung Chen, Peng Hsu Chen, Kuo Hao Ho, Chwen Ming Shih, Chih Ming Chou, Chia Hsiung ChengAbstract:Temozolomide (TMZ) is a first-line alkylating agent for glioblastoma multiforme (GBM). Clarifying the mechanisms inducing TMZ insensitivity may be helpful in improving its therapeutic effectiveness against GBM. Insulin-like growth factor (IGF)-1 signaling and micro (mi)RNAs are relevant in mediating GBM progression. However, their roles in desensitizing GBM cells to TMZ are still unclear. We aimed to identify IGF-1-mediated miRNA regulatory networks that elicit TMZ insensitivity for GBM. IGF-1 treatment attenuated TMZ cytotoxicity via WNT/beta-catenin signaling, but did not influence glioma cell growth. By miRNA array analyses, 93 upregulated and 148 downregulated miRNAs were identified in IGF-1-treated glioma cells. miR-513a-5p from the miR-513a-2 gene locus was upregulated by IGF-1-mediated phosphoinositide 3-kinase (PI3K) signaling. Its elevated levels were also observed in gliomas versus normal cells, in array data of The Cancer Genome Atlas (TCGA), and the GSE61710, GSE37366, and GSE41032 datasets. In addition, lower levels of neural precursor cell-expressed developmentally downregulated 4-like (NEDD4L), an E3 ubiquitin protein ligase that inhibits WNT signaling, were found in gliomas by analyzing cells, arrays, and RNA sequencing data of TCGA glioma patients. Furthermore, a negative correlation was identified between miR-513a-5p and NEDD4L in glioma. NEDD4L was also validated as a direct target gene of miR-513a-5p, and it was reduced by IGF-1 treatment. Overexpression of NEDD4L inhibited glioma cell viability and reversed IGF-1-repressed TMZ cytotoxicity. In contrast, miR-513a-5p significantly affected NEDD4L-inhibited WNT signaling and reduced TMZ cytotoxicity. These findings demonstrate a distinct role of IGF-1 signaling through miR-513a-5p-inhibited NEDD4L networks in influencing GBM's drug sensitivity to TMZ.
Daniela Rotin - One of the best experts on this subject based on the ideXlab platform.
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the ubiquitin ligase NEDD4L regulates the na k 2cl co transporter nkcc1 slc12a2 in the colon
Journal of Biological Chemistry, 2017Co-Authors: Chong Jiang, Hiroshi Kawabe, Daniela RotinAbstract:Abstract The ubiquitin ligase Nedd4-like (NEDD4L, or Nedd4-2) binds to and regulates stability of the epithelial Na+ channel (ENaC) in salt-absorbing epithelia in the kidney, lung, and other tissues. Its role in the distal colon, which also absorbs salt and fluid and expresses ENaC, is unknown. Using a conditional knock-out approach to knock out NEDD4L in mice intestinal epithelium (NEDD4Lf/f;Vil-CreERT2) we show here that NEDD4L depletion leads to a higher steady-state short circuit current (Isc) in mouse distal colon tissue relative to controls. This higher Isc was partially reduced by the addition of apical amiloride and strongly reduced by basolateral bumetanide as well as by depletion of basolateral Cl−, suggesting that Na+/K+/2Cl− (NKCC1/SLC12A2) co-transporter and ENaC are targets of NEDD4L in the colon. In accordance, NKCC1 (and γENaC) protein abundance in the colon of the NEDD4L knock-out animals was increased, indicating that NEDD4L normally suppresses these proteins. However, we did not observe co-immunoprecipitation between NEDD4L and NKCC1, suggesting that NEDD4L indirectly suppresses NKCC1 expression. Low salt diet resulted in a strong increase in β and γ (but not α) ENaC mRNA and protein expression and ENaC activity. Although salt restriction also increased NKCC1 protein and mRNA abundance, it did not lead to its elevated activity (Isc). These results identify NKCC1 as a novel target for NEDD4L-mediated down-regulation in vivo, which modulates ion and fluid transport in the distal colon together with ENaC.
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The Ubiquitin Ligase NEDD4L Regulates the Na/K/2Cl Co-transporter NKCC1/SLC12A2 in the Colon
Journal of Biological Chemistry, 2017Co-Authors: Chong Jiang, Hiroshi Kawabe, Daniela RotinAbstract:Abstract The ubiquitin ligase Nedd4-like (NEDD4L, or Nedd4-2) binds to and regulates stability of the epithelial Na+ channel (ENaC) in salt-absorbing epithelia in the kidney, lung, and other tissues. Its role in the distal colon, which also absorbs salt and fluid and expresses ENaC, is unknown. Using a conditional knock-out approach to knock out NEDD4L in mice intestinal epithelium (NEDD4Lf/f;Vil-CreERT2) we show here that NEDD4L depletion leads to a higher steady-state short circuit current (Isc) in mouse distal colon tissue relative to controls. This higher Isc was partially reduced by the addition of apical amiloride and strongly reduced by basolateral bumetanide as well as by depletion of basolateral Cl−, suggesting that Na+/K+/2Cl− (NKCC1/SLC12A2) co-transporter and ENaC are targets of NEDD4L in the colon. In accordance, NKCC1 (and γENaC) protein abundance in the colon of the NEDD4L knock-out animals was increased, indicating that NEDD4L normally suppresses these proteins. However, we did not observe co-immunoprecipitation between NEDD4L and NKCC1, suggesting that NEDD4L indirectly suppresses NKCC1 expression. Low salt diet resulted in a strong increase in β and γ (but not α) ENaC mRNA and protein expression and ENaC activity. Although salt restriction also increased NKCC1 protein and mRNA abundance, it did not lead to its elevated activity (Isc). These results identify NKCC1 as a novel target for NEDD4L-mediated down-regulation in vivo, which modulates ion and fluid transport in the distal colon together with ENaC.
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deletion of the ubiquitin ligase NEDD4L in lung epithelia causes cystic fibrosis like disease
Proceedings of the National Academy of Sciences of the United States of America, 2011Co-Authors: Toshihiro Kimura, Chong Jiang, Hiroshi Kawabe, Wenbo Zhang, Yunyan Xiang, Chen Lu, Michael W Salter, Nils Brose, Weiyang Lu, Daniela RotinAbstract:Cystic fibrosis is caused by impaired ion transport due to mutated cystic fibrosis transmembrane conductance regulator, accompanied by elevated activity of the amiloride-sensitive epithelial Na+ channel (ENaC). Here we show that knockout of the ubiquitin ligase NEDD4L (Nedd4-2) specifically in lung epithelia (surfactant protein C-expressing type II and Clara cells) causes cystic fibrosis-like lung disease, with airway mucus obstruction, goblet cell hyperplasia, massive inflammation, fibrosis, and death by three weeks of age. These effects of NEDD4L loss are likely caused by enhanced ENaC function, as reflected by increased ENaC protein levels, increased lung dryness at birth, amiloride-sensitive dehydration of lung explants, and elevated ENaC currents in primary alveolar type II cells analyzed by patch clamp recordings. Moreover, the lung defects were rescued with administration of amiloride into the lungs of young knockout pups via nasal instillation. Our results therefore suggest that the ubiquitin ligase NEDD4L can suppress the onset of cystic fibrosis symptoms by inhibiting ENaC in lung epithelia.
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defective regulation of the epithelial na channel by nedd4 in liddle s syndrome
Journal of Clinical Investigation, 1999Co-Authors: Hugues Abriel, Daniela Rotin, Johannes Loffing, John F Rebhun, Howard J Pratt, Laurent Schild, Jeandaniel Horisberger, Olivier StaubAbstract:Liddle's syndrome is an inherited form of hypertension linked to mutations in the epithelial Na+ channel (ENaC). ENaC is composed of three subunits (α, β, γ), each containing a COOH-terminal PY motif (xPPxY). Mutations causing Liddle's syndrome alter or delete the PY motifs of β- or γ-ENaC. We recently demonstrated that the ubiquitin–protein ligase Nedd4 binds these PY motifs and that ENaC is regulated by ubiquitination. Here, we investigate, using the Xenopus oocyte system, whether Nedd4 affects ENaC function. Overexpression of wild-type Nedd4, together with ENaC, inhibited channel activity, whereas a catalytically inactive Nedd4 stimulated it, likely by acting as a competitive antagonist to endogenous Nedd4. These effects were dependant on the PY motifs, because no Nedd4-mediated changes in channel activity were observed in ENaC lacking them. The effect of Nedd4 on ENaC missing only one PY motif (of β-ENaC), as originally described in patients with Liddle's syndrome, was intermediate. Changes were due entirely to alterations in ENaC numbers at the plasma membrane, as determined by surface binding and immunofluorescence. Our results demonstrate that Nedd4 is a negative regulator of ENaC and suggest that the loss of Nedd4 binding sites in ENaC observed in Liddle's syndrome may explain the increase in channel number at the cell surface, increased Na+ reabsorption by the distal nephron, and hence the hypertension. J. Clin. Invest. 103:667–673 (1999)
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Defective regulation of the epithelial Na+ channel by Nedd4 in Liddle's syndrome
Journal of Clinical Investigation, 1999Co-Authors: Hugues Abriel, Daniela Rotin, Johannes Loffing, John F Rebhun, Laurent Schild, Jeandaniel Horisberger, J. Howard Pratt, Olivier StaubAbstract:Liddle's syndrome is an inherited form of hypertension linked to mutations in the epithelial Na+ channel (ENaC). ENaC is composed of three subunits (α, β, γ), each containing a COOH-terminal PY motif (xPPxY). Mutations causing Liddle's syndrome alter or delete the PY motifs of β- or γ-ENaC. We recently demonstrated that the ubiquitin–protein ligase Nedd4 binds these PY motifs and that ENaC is regulated by ubiquitination. Here, we investigate, using the Xenopus oocyte system, whether Nedd4 affects ENaC function. Overexpression of wild-type Nedd4, together with ENaC, inhibited channel activity, whereas a catalytically inactive Nedd4 stimulated it, likely by acting as a competitive antagonist to endogenous Nedd4. These effects were dependant on the PY motifs, because no Nedd4-mediated changes in channel activity were observed in ENaC lacking them. The effect of Nedd4 on ENaC missing only one PY motif (of β-ENaC), as originally described in patients with Liddle's syndrome, was intermediate. Changes were due entirely to alterations in ENaC numbers at the plasma membrane, as determined by surface binding and immunofluorescence. Our results demonstrate that Nedd4 is a negative regulator of ENaC and suggest that the loss of Nedd4 binding sites in ENaC observed in Liddle's syndrome may explain the increase in channel number at the cell surface, increased Na+ reabsorption by the distal nephron, and hence the hypertension. J. Clin. Invest. 103:667–673 (1999)
Yeguang Chen - One of the best experts on this subject based on the ideXlab platform.
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NEDD4L regulates convergent extension movements in xenopus embryos via disheveled mediated non canonical wnt signaling
Developmental Biology, 2014Co-Authors: Yan Zhang, Yi Ding, Yeguang ChenAbstract:During the early vertebrate body plan formation, convergent extension (CE) of dorsal mesoderm and neurectoderm is coordinated by the evolutionarily conserved non-canonical Wnt/PCP signaling. Disheveled (Dvl), a key mediator of Wnt/PCP signaling, is essential for the medial–lateral polarity formation in the cells undergoing convergent extension movements. NEDD4L, a highly conserved HECT type E3 ligase, has been reported to regulate the stability of multiple substrates including Dvl2. Here we demonstrate that NEDD4L is required for the cellular polarity formation and convergent extension in the early Xenopus embryos. Depletion of NEDD4L in early Xenopus embryos results in the loss of mediolateral polarity of the convergent-extending mesoderm cells and the shortened body axis, resembling those defects caused by the disruption of non-canonical Wnt signaling. Depletion of xNEDD4L also blocks the elongation of the animal explants in response to endogenous mesoderm inducing signals and partially compromises the expression of Brachyury. Importantly, reducing Dvl2 expression can largely rescue the cellular polarity and convergent extension defects in NEDD4L-depleted embryos and explants. Together with the data that NEDD4L reduces Dvl2 protein expression in the frog embryos, our findings suggest that regulation of Dvl protein levels by NEDD4L is essential for convergent extension during early Xenopus embryogenesis.
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hect domain containing e3 ubiquitin ligase NEDD4L negatively regulates wnt signaling by targeting dishevelled for proteasomal degradation
Journal of Biological Chemistry, 2013Co-Authors: Yi Ding, Yan Zhang, Chao Xu, Yeguang ChenAbstract:Wnt signaling plays a pivotal role in embryogenesis and tissue homeostasis. Dishevelled (Dvl) is a central mediator for both Wnt/β-catenin and Wnt/planar cell polarity pathways. NEDD4L, an E3 ubiquitin ligase, has been shown to regulate ion channel activity, cell signaling, and cell polarity. Here, we report a novel role of NEDD4L in the regulation of Wnt signaling. NEDD4L induces Dvl2 polyubiquitination and targets Dvl2 for proteasomal degradation. Interestingly, the NEDD4L-mediated ubiquitination of Dvl2 is Lys-6, Lys-27, and Lys-29 linked but not typical Lys-48-linked ubiquitination. Consistent with the role of Dvl in both Wnt/β-catenin and Wnt/planar cell polarity signaling, NEDD4L regulates the cellular β-catenin level and Rac1, RhoA, and JNK activities. We have further identified a hierarchical regulation that Wnt5a induces JNK-mediated phosphorylation of NEDD4L, which in turn promotes its ability to degrade Dvl2. Finally, we show that NEDD4L inhibits Dvl2-induced axis duplication in Xenopus embryos. Our work thus demonstrates that NEDD4L is a negative feedback regulator of Wnt signaling.
Qien Wang - One of the best experts on this subject based on the ideXlab platform.
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mir 93 promotes tgf β induced epithelial to mesenchymal transition through downregulation of NEDD4L in lung cancer cells
Tumor Biology, 2016Co-Authors: Meihua Qu, Amit Srivastava, Qien WangAbstract:The level of microRNA-93 (miR-93) in tumors has been recently reported to be negatively correlated with survival of lung cancer patients. Considering that the most devastating aspect of lung cancer is metastasis, which can be promoted by transforming growth factor-β (TGF-β)-induced epithelial-to-mesenchymal transition (EMT), we sought to determine whether miR-93 is involved in this process. Here, we report that a previously unidentified target of miR-93, neural precursor cell expressed developmentally downregulated gene 4-like (NEDD4L), is able to mediate TGF-β-mediated EMT in lung cancer cells. miR-93 binds directly to the 3′-UTR of the NEDD4L messenger RNA (mRNA), leading to a downregulation of NEDD4L expression at the protein level. We next demonstrated that the downregulation of NEDD4L enhanced, while overexpression of NEDD4L reduced TGF-β signaling, reflected by increased phosphorylation of SMAD2 in the lung cancer cell line after TGF-β treatment. Furthermore, overexpression of miR-93 in lung cancer cells promoted TGF-β-induced EMT through downregulation of NEDD4L. The analysis of publicly available gene expression array datasets indicates that low NEDD4L expression correlates with poor outcomes among patients with lung cancer, further supporting the oncogenic role of miR-93 in lung tumorigenesis and metastasis.
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ddb2 modulates tgf β signal transduction in human ovarian cancer cells by downregulating NEDD4L
Nucleic Acids Research, 2015Co-Authors: Ran Zhao, Amit Srivastava, Qien Wang, Xiaoli Zhang, Jinshan He, Jianhua Yu, Altaf A WaniAbstract:The expression of DNA damage-binding protein 2 (DDB2) has been linked to the prognosis of ovarian cancer and its underlying transcription regulatory function was proposed to contribute to the favorable treatment outcome. By applying gene microarray analysis, we discovered neural precursor cell expressed, developmentally downregulated 4-Like (NEDD4L) as a previously unidentified downstream gene regulated by DDB2. Mechanistic investigation demonstrated that DDB2 can bind to the promoter region of NEDD4L and recruit enhancer of zeste homolog 2 histone methyltransferase to repress NEDD4L transcription by enhancing histone H3 lysine 27 trimethylation (H3K27me3) at the NEDD4L promoter. Given that NEDD4L plays an important role in constraining transforming growth factor β signaling by targeting activated Smad2/Smad3 for degradation, we investigated the role of DDB2 in the regulation of TGF-β signaling in ovarian cancer cells. Our data indicate that DDB2 enhances TGF-β signal transduction and increases the responsiveness of ovarian cancer cells to TGF-β-induced growth inhibition. The study has uncovered an unappreciated regulatory mode that hinges on the interaction between DDB2 and NEDD4L in human ovarian cancer cells. The novel mechanism proposes the DDB2-mediated fine-tuning of TGF-β signaling and its downstream effects that impinge upon tumor growth in ovarian cancers.
