HDAC-mediated control of ERK- and PI3K-dependent TGF-β-induced extracellular matrix-regulating genes

Histone deacetylases (HDACs) regulate the acetylation of histones in the control of gene expression. Many non-histone proteins are also targeted for acetylation, including TGF-β signalling pathway components such as Smad2, Smad3 and Smad7. Our studies in mouse C3H10T1/2 fibroblasts suggested that a number of TGF-β-induced genes that regulate matrix turnover are selectively regulated by HDACs. Blockade of HDAC activity with trichostatin A (TSA) abrogated the induction of a disintegrin and metalloproteinase 12 (Adam12) and tissue inhibitor of metalloproteinases-1 (Timp-1) genes by TGF-β, whereas plasminogen activator inhibitor-1 (Pai-1) expression was unaffected. Analysis of the activation of cell signalling pathways demonstrated that TGF-β induced robust ERK and PI3K activation with delayed kinetics compared to the phosphorylation of Smads. The TGF-β induction of Adam12 and Timp-1 was dependent on such non-Smad signalling pathways and, importantly, HDAC inhibitors completely blocked their activation without affecting Smad signalling. Analysis of TGF-β-induced Adam12 and Timp-1 expression and ERK/PI3K signalling in the presence of semi-selective HDAC inhibitors valproic acid, MS-275 and apicidin implicated a role for class I HDACs. Furthermore, depletion of HDAC3 by RNA interference significantly down-regulated TGF-β-induced Adam12 and Timp-1 expression without modulating Pai-1 expression. Correlating with the effect of HDAC inhibitors, depletion of HDAC3 also blocked the activation of ERK and PI3K by TGF-β. Collectively, these data confirm that HDACs, and in particular HDAC3, are required for activation of the ERK and PI3K signalling pathways by TGF-β and for the subsequent gene induction dependent on these signalling pathways.     

TGF-β/Smads信号转导通路的研究进展

转化生长因子（TGF）-β超家族成员的重要生物学功能正日益引起人们的重视。受体介导的胞内信号转导研究近年有较大进展,特别是Smads蛋白介导的信号转导通路为阐明TGF-β超家族的作用机理提供了一条重要线索。TGF-β/Smads信号的转导受到机体严密的调控,并与其他信号通路存在着广泛的交叉对话效应。综述了对TGF-β/Smads信号转导通路的机制、调控,及其在维持机体正常生理功能和疾病发生中的作用的研究进展。     

Apocynin inhibits the upregulation of TGF-β1 expression and ROS production induced by TGF-β in skeletal muscle cells

BackgroundPure apocynin, which can be traditionally isolated and purified from several plant species such as Picrorhiza kurroa Royle ex Benth (Scrophulariaceae), acts as an inhibitor of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activity inhibiting its production of reactive oxygen species (ROS). Transforming growth factor type beta 1 (TGF-β1) is a growth factor that produces inhibition of myogenesis, diminution of regeneration and induction of atrophy in skeletal muscle. The typical signalling that is activated by TGF-β involves the Smad pathway.PurposeTo evaluate the effect of TGF-β and the effect of apocynin on TGF-β1 expression in skeletal muscle cells.Study designControlled laboratory study. In vitro assays were performed with C₂C₁₂ cells incubated with TGF-β1 in presence or absence of apocynin (NOX inhibitor), SB525334 (TGF-β-receptor I inhibitor), or chelerythrine (PKC inhibitor).MethodsTGF-β1 and atrogin-1 expression was evaluated by RT-qPCR and/or ELISA; Smad3 phosphorylation by western blot; Smad4 nuclear translocation by indirect immunofluorescence; and ROS levels by DCF probe fluorescent measurements.ResultsWe show that myoblasts respond to TGF-β1 by increasing its own gene expression in a time- and dose-dependent fashion which was abolished by SB525334 and siRNA for Smad2/3. TGF-β1 also induced ROS. Remarkably, apocynin inhibited the TGF-β1 induced ROS as well as the autoinduction of TGF-β1 gene expression. We also show that TGF-β-induced ROS production and TGF-β1 expression require PKC activity as indicated by the inhibition using chelerythrine.ConclusionThese results strongly suggest that TGF-β induces its own expression through a TGF-β-receptor/Smad-dependent mechanism and apocynin is able to inhibit this process, suggesting that requires NOX-induced ROS in skeletal muscle cells.     

SARA is dispensable for functional TGF-β signaling

Smad anchor for receptor activation (SARA or ZFYVE9) has been proposed to mediate transforming growth factor β (TGF-β) signaling by direct interaction with the non-activated Smad proteins and the TGF-β receptors; however, these findings are controversial. We demonstrate no correlation between SARA expression and the levels of TGF-β-induced phosphorylation of Smads in various B-cell lymphomas. Moreover, knockdown of SARA in HeLa cells did not interfere with TGF-β-induced Smad activation, Smad nuclear translocation, or induction of TGF-β target genes. Various R-Smads and TGF-β receptors did not co-immunoprecipitate with SARA. Collectively, our results demonstrate that SARA is dispensable for functional TGF-β-mediated signaling.     

Effects of SARA on Oxygen–Glucose Deprivation in PC12 Cell Line

Ischemic stroke is a major composition of cerebrovascular disease, seriously threatening to human health in the world. Activin A (ActA), belonging to transforming growth factor-beta (TGF-β) super family, plays an important role in the hypoxic-ischemic brain injury through ActA/Smads pathway. While as an essential phosphorylation assistor in TGF-β signaling, the functions and mechanisms of smad anchor for receptor activation (SARA) in ischemic brain injury remain poorly understood. To solve this problem and explore the pathological processes of ischemic stroke, we used an Oxygen–Glucose deprivation (OGD) model in nerve growth factor-induced differentiated rattus PC12 pheochromocytoma cells and down regulated the expressions of SARA by RNA interference technology. Our results showed that the repression of SARA before OGD exposure reduced the expressions of Smad2, 3, 4 mRNA and the phosphorylation rate of Smad2 protein, but it did not affect the mRNA expressions of Smad7. After OGD treatment, ActA/Smads pathway was activated and the expression of SARA in the SARA pre-repression group was significantly up-regulated. The pre-repression of SARA increased the sensitivities of nerve-like cells to OGD damage. Moreover, the mRNA expression of Smad7 which was supposed to participate in the negative feedback of ActA/Smads pathway was also elevated due to OGD injury. Taken together, these results suggest a positive role of SARA in assisting the phosphorylation of Smad2 and maintaining the neuron protective effect of ActA/Smads pathway.     

TGF-β/Smads通路转导蛋白在心力衰竭发病中的研究进展

转化生长因子-β（transforming growth factor-β,TGF-β）作用复杂,广泛参与哺乳动物的各种病理生理过程如影响细胞的增殖分化、参与心力衰竭发展。TGF-β信号通路关键的信号传导分子为胞浆蛋白Smads。TGF-β诱导分化心肌成纤维细胞,刺激胶原蛋白等细胞间质成分的合成,促进细胞间质的沉积和抑制弹性蛋白酶等的分泌,刺激蛋白酶抑制剂的表达,从而抑制Ⅰ型胶原成分的降解,促进心肌纤维化发展,进一步了解TGF-β/Smads通路转导蛋白的作用及机制,通过抑制TGF-β/Smads通路转导蛋白作为治疗新靶点为最终防止心力衰竭提供了新的思路。     

逆转肾小管上皮细胞转分化的Smad信号转导机制

肾小管上皮细胞转分化(tubular epithelial to menchymal transdifferentiation,EMT)是肾小管间质纤维化的重要病理机制之一。致纤维化细胞因子TGF-β通过几种信号转导途径调节EMT,其中TGF-β/Smads信号通路发挥核心作用。目前研究表明,Smad7、HGF、BMP-7等可通过调控Smads信号通路而逆转EMT,这为肾间质纤维化的防治提供了新的思路。该文主要介绍TGF-β/Smads信号通路在EMT发生的作用,以及Smad7、SnoN、HGF、BMP-7等分子是如何通过抑制Smads信号通路而发挥逆转EMT作用的。