A novel transcriptional factor with Ser/Thr kinase activity involved in the transforming growth factor (TGF)-beta signalling pathway.
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ABSTRACT:
SUBMITTER: Ohta S
PROVIDER: S-EPMC1221266 | biostudies-other | 2000 Sep
REPOSITORIES: biostudies-other
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Project description:Transforming growth factor-β (TGF-β) signaling regulates cell proliferation and differentiation, which contributes to development and disease. Upon binding TGF-β, the type I receptor (TGFBRI) binds TGFBRII, leading to the activation of the transcription factors SMAD2 and SMAD3. Using an RNA interference screen of the human kinome and a live-cell reporter for TGFBR activity, we identified the kinase BUB1 (budding uninhibited by benzimidazoles-1) as a key mediator of TGF-β signaling. BUB1 interacted with TGFBRI in the presence of TGF-β and promoted the heterodimerization of TGFBRI and TGFBRII. Additionally, BUB1 interacted with TGFBRII, suggesting the formation of a ternary complex. Knocking down BUB1 prevented the recruitment of SMAD3 to the receptor complex, the phosphorylation of SMAD2 and SMAD3 and their interaction with SMAD4, SMAD-dependent transcription, and TGF-β-mediated changes in cellular phenotype including epithelial-mesenchymal transition (EMT), migration, and invasion. Knockdown of BUB1 also impaired noncanonical TGF-β signaling mediated by the kinases AKT and p38 MAPK (mitogen-activated protein kinase). The ability of BUB1 to promote TGF-β signaling depended on the kinase activity of BUB1. A small-molecule inhibitor of the kinase activity of BUB1 (2OH-BNPP1) and a kinase-deficient mutant of BUB1 suppressed TGF-β signaling and formation of the ternary complex in various normal and cancer cell lines. 2OH-BNPP1 administration to mice bearing lung carcinoma xenografts reduced the amount of phosphorylated SMAD2 in tumor tissue. These findings indicated that BUB1 functions as a kinase in the TGF-β pathway in a role beyond its established function in cell cycle regulation and chromosome cohesion.
| S-EPMC4440544 | biostudies-literature
Project description:Megalin/LRP2 is a receptor that plays important roles in the physiology of several organs, such as kidney, lung, intestine, and gallbladder and also in the physiology of the nervous system. Megalin expression is reduced in diseases associated with fibrosis, including diabetic nephropathy, hepatic fibrosis and cholelithiasis, as well as in some breast and prostate cancers. One of the hallmarks of these conditions is the presence of the cytokine transforming growth factor beta (TGF-ß). Although TGF-ß has been implicated in the reduction of megalin levels, the molecular mechanism underlying this regulation is not well understood. Here, we show that treatment of two epithelial cell lines (from kidney and gallbladder) with TGF-ß1 is associated with decreased megalin mRNA and protein levels, and that these effects are reversed by inhibiting the TGF-ß1 type I receptor (TGF-ßRI). Based on in silico analyses, the two SMAD-binding elements (SBEs) in the megalin promoter are located at positions -57 and -605. Site-directed mutagenesis of the SBEs and chromatin immunoprecipitation (ChIP) experiments revealed that SMAD2/3 transcription factors interact with SBEs. Both the presence of SMAD2/3 and intact SBEs were associated with repression of the megalin promoter, in the absence as well in the presence of TGF-ß1. Also, reduced megalin expression and promoter activation triggered by high concentration of albumin are dependent on the expression of SMAD2/3. Interestingly, the histone deacetylase inhibitor Trichostatin A (TSA), which induces megalin expression, reduced the effects of TGF-ß1 on megalin mRNA levels. These data show the significance of TGF-ß and the SMAD2/3 signalling pathway in the regulation of megalin and explain the decreased megalin levels observed under conditions in which TGF-ß is upregulated, including fibrosis-associated diseases and cancer.
| S-EPMC6532859 | biostudies-literature
Project description:BACKGROUND F-box protein 32 (FBXO32) (also known as atrogin-1), a member of the F-box protein family, was recently shown to be a transforming growth factor beta (TGF-?)/Smad4 target gene involved in regulating cell survival. It can be transcriptionally silenced by epigenetic mechanisms in some cancers, but its role in colorectal carcinoma (CRC) is unclear. We investigated the role of FBXO32 in CRC and determined its prognostic significance. MATERIAL AND METHODS We used real-time quantitative PCR, Western blot, and immunohistochemistry to elucidate the role of FBXO32 in clinical specimens and primary CRC cell lines. Differences in patient survival were determined by the Kaplan-Meier method and log-rank test. RESULTS We found that the FBXO32 and SMAD4 levels were higher in normal tissues than in CRC tissues, but PAI-1 and VEGF levels showed the opposite pattern. The expressions of FBXO32 and SMAD4 were related to clinicopathological parameters in CRC. Kaplan-Meier analyses showed that the 5-year overall survival of the low-FBXO32 expression group was significantly shorter than that of the high-FBXO32 expression group (p=0.010). CONCLUSIONS The fbxo32 gene is a novel tumor suppressor that inhibits CRC progression by inducing differentiation. Elevated expression of FBXO32 predicts longer survival in CRC patients.
| S-EPMC5829536 | biostudies-literature
Project description:Patients with congenital adrenal hyperplasia (CAH) with tenascin-X deficiency (CAH-X syndrome) have both endocrine imbalances and characteristic Ehlers Danlos syndrome phenotypes. Unlike other subtypes, tenascin-X-related Ehlers Danlos syndrome is caused by an extracellular matrix protein deficiency rather than a defect in fibrillar collagen or a collagen-modifying enzyme, and the understanding of the disease mechanisms is limited. We hypothesized that transforming growth factor-β pathway dysregulation may, in part, be responsible for connective tissue phenotypes observed in CAH-X, due to this pathway's known role in connective tissue disorders. Fibroblasts and direct tissue from human skin biopsies from CAH-X probands and age- and sex-matched controls were screened for transforming growth factor-β biomarkers known to be dysregulated in other hereditary disorders of connective tissue. In CAH-X fibroblast lines and dermal tissue, pSmad1/5/8 was significantly upregulated compared to controls, suggesting involvement of the bone morphogenetic protein pathway. Additionally, CAH-X samples compared to controls exhibited significant increases in fibroblast-secreted TGF-β3, a cytokine important in secondary palatal development, and in plasma TGF-β2, a cytokine involved in cardiac function and development, as well as palatogenesis. Finally, MMP-13, a matrix metalloproteinase important in secondary palate formation and tissue remodeling, had significantly increased mRNA and protein expression in CAH-X fibroblasts and direct tissue. Collectively, these results demonstrate that patients with CAH-X syndrome exhibit increased expression of several transforming growth factor-β biomarkers and provide a novel link between this signaling pathway and the connective tissue dysplasia phenotypes associated with tenascin-X deficiency.
| S-EPMC3972366 | biostudies-literature
Project description:CCN5 is a member of the CCN (connective tissue growth factor/cysteine-rich 61/nephroblastoma overexpressed) family and was identified as an estrogen-inducible gene in estrogen receptor-positive cell lines. However, the role of CCN5 in breast carcinogenesis remains unclear. We report here that the CCN5 protein is localized mostly in the cytoplasm and in part in the nucleus of human tumor breast tissue. Using a heterologous transcription assay, we demonstrate that CCN5 can act as a transcriptional repressor presumably through association with histone deacetylase 1 (HDAC1). Microarray gene expression analysis showed that CCN5 represses expression of genes associated with epithelial-mesenchymal transition (EMT) as well as expression of key components of the transforming growth factor β (TGF-β) signaling pathway, prominent among them TGF-βRII receptor. We show that CCN5 is recruited to the TGF-βRII promoter, thereby providing a mechanism by which CCN5 restricts transcription of the TGF-βRII gene. Consistent with this finding, CCN5, we found, functions to suppress TGF-β-induced transcriptional responses and invasion that is concomitant with EMT. Thus, our data uncovered CCN5 as a novel transcriptional repressor that plays an important role in regulating tumor progression functioning, at least in part, by inhibiting the expression of genes involved in the TGF-β signaling cascade that is known to promote EMT.
| S-EPMC3135290 | biostudies-literature
Project description:BackgroundChronic fatigue syndrome (CFS) is a prevalent and disabling condition among adolescent. The disease mechanisms are unknown. Previous studies have suggested elevated plasma levels of several cytokines, but a recent meta-analysis of 38 articles found that of 77 different cytokines measured in plasma, transforming growth factor beta (TGF-β) was the only one that was elevated in patients compared to controls in a sufficient number of articles. In the present study we therefore compared the plasma levels of the three TGF-β isoforms in adolescent CFS patients and healthy controls. In addition, the study explored associations between TGF-β levels, neuroendocrine markers, clinical markers and differentially expressed genes within the CFS group.MethodsCFS patients aged 12-18 years (n = 120) were recruited nation-wide to a single referral center as part of the NorCAPITAL project (ClinicalTrials ID: NCT01040429). A broad case definition of CFS was applied, requiring 3 months of unexplained, disabling chronic/relapsing fatigue of new onset, whereas no accompanying symptoms were necessary. Healthy controls (n = 68) were recruited from local schools. The three isoforms of TGF-β (TGF-β1, TGF-β2, TGF-β3) were assayed using multiplex technology. Neuroendocrine markers encompassed plasma and urine levels of catecholamines and cortisol, as well as heart rate variability indices. Clinical markers consisted of questionnaire scores for symptoms of post-exertional malaise, inflammation, fatigue, depression and trait anxiety, as well as activity recordings. Whole blood gene expression was assessed by RNA sequencing in a subgroup of patients (n = 29) and controls (n = 18).ResultsPlasma levels of all three isoforms of TGF-β were equal in the CFS patients and the healthy controls. Subgrouping according to the Fukuda and Canada 2003 criteria of CFS did not reveal differential results. Within the CFS group, all isoforms of TGF-β were associated with plasma cortisol, urine norepinephrine and urine epinephrine, and this association pattern was related to fatigue score. Also, TGF-β3 was related to expression of the B cell annotated genes TNFRSF13C and CXCR5.ConclusionsPlasma levels of all TGF-β isoforms were not altered in adolescent CFS. However, the TGF-β isoforms were associated with neuroendocrine markers, an association related to fatigue score. Furthermore, TGF-β3 might partly mediate an association between plasma cortisol and B cell gene expression. Trial registration Clinical Trials NCT01040429.
| S-EPMC5716371 | biostudies-literature
Project description:Appropriate growth and development of the endometrium across the menstrual cycle is key for a woman's quality of life and reproductive well-being. Recurrent pregnancy loss (RPL) and heavy menstrual bleeding (HMB) affect a significant proportion of the female population worldwide. These endometrial pathologies have a significant impact on a woman's quality of life as well as placing a high economic burden on a country's health service. An underlying cause for both conditions is unknown in approximately 50% of cases. Previous research has demonstrated that aberrant endometrial vascular maturation is associated with both RPL and HMB, where it is increased in RPL but reduced in HMB. TGFβ1 is one of the key growth factors that regulate vascular maturation, by inducing phenotypic switching of vascular smooth muscle cells (VSMCs) from a synthetic phenotype to a more contractile one. Our previous data demonstrated an increase in TGFβ1 in the endometrium of RPL, while others have shown a decrease in women with HMB. However, TGFβ1 bioavailability is tightly controlled, and we therefore sought to perform an extensive immunohistochemical analysis of different components in the pathway in the endometrium of normal controls, women with HMB or RPL. In addition, two in vitro models were used to examine the role of TGFβ1 in endometrial vascular maturation and endothelial cell (EC):VSMC association. Taken all together, the immunohistochemical data suggest a decrease in bioavailability, receptor binding capacity, and signaling in the endometrium of women with HMB compared with controls. In contrast, there is an increase in the bioavailability of active TGFβ1 in the endometrium of women with RPL compared with controls. Endometrial explants cultured in TGFβ1 had an increase in the number of vessels associated with contractile VSMC markers, although the total number of vessels did not increase. In addition, TGFβ1 increased EC:VSMC association in an in vitro model. In conclusion, TGFβ1 is a key regulator of endometrial vascular maturation and could be considered as a therapeutic target for women suffering from HMB and/or RPL.
| S-EPMC8063037 | biostudies-literature
Project description:Chronic renal failure (CRF), also known as chronic kidney disease (CKD), is a common renal disorder characterized by gradual kidney dysfunction. Molecular dissection reveals that transforming growth factor beta (TGF-β) plays a central role in the pathogenesis of CRF. However, the mechanism underlying TGF-β upregulation has not been demonstrated. Here, we verified that the elevated level of TGF-β was associated with the severity of CRF stages and the activation of TGF-β-mediated signaling in 120 renal biopsies from CRF patients. By analyzing the promoter region of the TGFB1 gene, we identified one AP-1 (activator protein 1) and four NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) binding sites. Knockdown of two AP-1 subunits (c-Jun and c-FOS) or blockage of AP-1 signaling with two inhibitors T-5224 and SR11302 could cause the downregulation of TGFB1, whereas knockdown of two NF-κB subunits (p65 and p50) or blockage of NF-κB signaling with two inhibitors TPCA1 and BOT-64 could not change the expression of TGFB1. Using mass spectrometry and coimmunoprecipitation analyses, we found that both c-Jun and c-FOS formed a complex with CtBP2 (C-terminal binding protein 2) and histone acetyltransferase p300. Our in vitro data demonstrated that induction of CtBP2 by recombinant IL-1β (interleukin-1 beta) led to the upregulation of TGFB1 and the activation of TGF-β downstream signaling, while knockdown of CtBP2 resulted in the reversed effects. Using chromatin immunoprecipitation assays, we revealed that the CtBP2-p300-AP1 complex specifically bound to the promoter of TGFB and that knockdown or blockage of CtBP2 significantly decreased the occupancies of the p300 and AP-1 subunits. Our results support a model in which the CtBP2-p300-AP1 transcriptional complex activates the expression of TGFB1, increasing its production and extracellular secretion. The secreted TGF-β binds to its receptors and initiates downstream signaling.
| S-EPMC6949151 | biostudies-literature
Project description:Preeclampsia (PE), a condition during pregnancy that involves high blood pressure and proteinuria, is potentially fatal to both mother and child. PE currently has no known etiology or cure but has been tied to poor placental trophoblast cell migration. Increased levels of the toxic metal cadmium (Cd) have been associated with increased risk of developing PE, as well as miRNA-associated regulation of the transforming growth factorbeta (TGF-β) pathway. Signal reprogramming of the TGF-β pathway via epigenetic mechanisms is hypothesized to modify placental trophoblast function. In the present study we investigated the role of increased and decreased signaling of the TGF-β pathway in relation to Cd-induced reduction in cellular migration in JEG3 trophoblast cells. Furthermore, the role of a miR-26a as a molecular mediator of placental trophoblast migration was confirmed. The results demonstrate that increased expression of miR-26a and decreased signaling of the TGF-β pathway increase placental cell migration. These findings have relevance for mechanistic understanding of the underpinnings of poor placentation associated with PE.
| S-EPMC5656529 | biostudies-literature
Project description:We have described previously the use of microarray technology to identify novel target genes of TGF-beta (transforming growth factor-beta) signalling in mouse embryo fibroblasts deficient in Smad2 or Smad3 [Yang, Piek, Zavadil, Liang, Xie, Heyer, Pavlidis, Kucherlapati, Roberts and Böttinger (2003) Proc. Natl. Acad. Sci. U.S.A. 100, 10269-10274]. Among the TGF-beta target genes identified, a novel gene with sequence homology to members of the Ras superfamily was identified, which we have designated as RLP (Ras-like protein). RLP is a Smad3-dependent immediate-early TGF-beta target gene, its expression being induced within 45 min. Bone morphogenetic proteins also induce expression of RLP, whereas epidermal growth factor and phorbol ester PMA suppress TGF-beta-induced expression of RLP. Northern-blot analysis revealed that RLP was strongly expressed in heart, brain and kidney, and below the detection level in spleen and skeletal muscles. At the protein level, RLP is approx. 30% homologous with members of the Ras superfamily, particularly in domains characteristic for small GTPases. However, compared with prototypic Ras, RLP contains a modified P-loop, lacks the consensus G2 loop and the C-terminal prenylation site and harbours amino acid substitutions at positions that render prototypic Ras oncogenic. However, RLP does not have transforming activity, does not affect phosphorylation of mitogen-activated protein kinase and is unable to bind GTP or GDP. RLP was found to associate with certain subtypes of the TGF-beta receptor family, raising the possibility that RLP plays a role in TGF-beta signal transduction. Although RLP did not interact with Smads and did not affect TGF-beta receptor-induced Smad2 phosphorylation, it inhibited TGF-beta-induced transcriptional reporter activation, suggesting that it is a novel negative regulator of TGF-beta signalling.
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