Serum response factor

Serum response factor (SRF) transcriptionally regulates expression of contractile genes in smooth muscle cells (SMC). Lack or decrease of SRF is directly linked to a phenotypic change of SMC, leading to hypomotility of smooth muscle in the gastrointestinal (GI) tract. However, the molecular mechanism behind SRF-induced hypomotility in GI smooth muscle is largely unknown. We describe here how SRF plays a functional role in the regulation of the SMC contractility via myotonic dystrophy protein kinase (DMPK) and L-type calcium channel CACNA1C. GI SMC expressed Dmpk and Cacna1c genes into multiple alternative transcriptional isoforms. Deficiency of SRF in SMC of Srf knockout (KO) mice led to reduction of SRF-dependent DMPK, which down-regulated the expression of CACNA1C. Reduction of CACNA1C in KO SMC not only decreased intracellular Ca2+ spikes but also disrupted their coupling between cells resulting in decreased contractility. The role of SRF in the regulation of SMC phenotype and fu...

Epithelial-mesenchymal-myofibroblast transition (EMT), a key feature in organ fibrosis, is regulated by the state of intercellular contacts. Our recent studies have shown that an initial injury of cell–cell junctions is a prerequisite for transforming growth factor-β1 (TGF-β1)-induced transdifferentiation of kidney tubular cells into α-smooth muscle actin (SMA)–expressing myofibroblasts. Here we analyzed the underlying contact-dependent mechanisms. Ca2+ removal–induced disruption of intercellular junctions provoked Rho/Rho kinase (ROK)-mediated myosin light chain (MLC) phosphorylation and Rho/ROK-dependent SMA promoter activation. Importantly, myosin-based contractility itself played a causal role, because the myosin ATPase inhibitor blebbistatin or a nonphosphorylatable, dominant negative MLC (DN-MLC) abolished the contact disruption-triggered SMA promoter activation, eliminated the synergy between contact injury and TGF-β1, and suppressed SMA expression. To explore the responsible...

Regulation of gene transcription in vascular smooth muscle cells (VSMCs) by serum response factor (SRF) plays a crucial role in vascular development and in the pathophysiology of vascular diseases. Nevertheless, the regulation of specific genes by SRF in vascular diseases is poorly understood. Therefore, we investigated the regulation of smooth muscle myosin light chain kinase (smMLCK) by using spontaneously hypertensive rats (SHR) as an experimental model. We found that smMLCK expression in blood vessels increases during the development of hypertension and is always greater in blood vessels from SHR compared with normotensive rats. Analysis of the DNA sequences of the promoters isolated from SHR and normotensive rats revealed that SHR contain a 12-base pair insertion adjacent to the CArG box. This insertion increases SRF binding to the CArG box and positively regulates SRF-dependent promoter activity. The increase in smMLCK expression was blocked by dominant-negative SRF, dominant-...

Rho family GTPases Rac and Cdc42 are pivotal regulators of apoptosis in multiple cell types. However, little is known about the mechanism by which these GTPases are regulated in response to apoptotic stimuli. Here, we demonstrate that TIAM1, a Rac-specific guanine nucleotide exchange factor, is cleaved by caspases during apoptosis. TIAM1 cleavage occurs in multiple cell lines in response to diverse apoptotic stimuli such as ceramide, Fas, and serum deprivation. Processing occurs at residue 993 of TIAM1 and removes the NH2-terminal of TIAM’s two pleckstrin homology domains, leaving a stable fragment containing the Dbl homology and COOH-terminal pleckstrin homology domains. This leads to functional inactivation of TIAM1, as determined by failure of the cleavage product to stimulate GTP loading of Rac in vivo. Furthermore, this product is defective in signaling to two independent Rac effectors, c-Jun NH2-terminal kinase and serum response factor. Finally, we demonstrate that in cells t...

Serum response factor (SRF) is at the confluence of multiple signaling pathways controlling the transcription of immediate-early response genes and muscle-specific genes. There are active SRF target sequences in more than 50 genes expressed in the three muscle lineages including normal and diseased hearts. However, the role of SRF in heart formation has not been addressed in vivo thus far due to the early requirement of SRF for mesoderm formation. We have generated a conditional mutant of SRF by using Cre-LoxP strategy that will be extremely useful to study the role of SRF in embryonic and postnatal cardiac functions, as well as in other tissues. This report shows that heart-specific deletion of SRF in the embryo by using a new βMHC-Cre transgenic mouse line results in lethal cardiac defects between embryonic day 10.5 (E10.5) and E13.5, as evidenced by abnormally thin myocardium, dilated cardiac chambers, poor trabeculation, and a disorganized interventricular septum. At E9.5, we fo...

Regulation of skeletal muscle determination and differentiation in vertebrates centers on a core regulatory network which is composed of two families of transcription factors, the MyoD group basic helix-loop-helix (bHLH) muscle regulatory factors (MRFs) and the myocyte enhancer factor 2 (MEF2) group of MADS-box regulators. Members of this network interact with each other genetically and physically, and together they cooperate to positively regulate transcription of downstream muscle-specific differentiation genes. During development, the myogenic network can be activated or repressed in response to patterning signals, some of which have recently been identified. Once activated, the powerful myogenic activity of the core network can be modulated and held in check by a remarkably large group of negative regulators that operate on network components by diverse mechanisms. Recent discoveries highlight extensive parallels between myogenesis and peripheral neurogenesis in the structures o...

Serum response factor (SRF) regulates certain microRNAs that play a role in cardiac and skeletal muscle development. However, the role of SRF in the regulation of microRNA expression and microRNA biogenesis in cardiac hypertrophy has not been well established. In this report, we employed two distinct transgenic mouse models to study the impact of SRF on cardiac microRNA expression and microRNA biogenesis. Cardiac-specific overexpression of SRF (SRF-Tg) led to altered expression of a number of microRNAs. Interestingly, downregulation of miR-1, miR-133a and upregulation of miR-21 occurred by 7 days of age in these mice, long before the onset of cardiac hypertrophy, suggesting that SRF overexpression impacted the expression of microRNAs which contribute to cardiac hypertrophy. Reducing cardiac SRF level using the antisense-SRF transgenic approach (Anti-SRF-Tg) resulted in the expression of miR-1, miR-133a and miR-21 in the opposite direction. Furthermore, we observed that SRF regulates microRNA biogenesis, specifically the transcription of pri-microRNA, thereby affecting the mature microRNA level. The mir-21 promoter sequence is conserved among mouse, rat and human; one SRF binding site was found to be in the mir-21 proximal promoter region of all three species. The mir-21 gene is regulated by SRF and its cofactors, including myocardin and p49/Strap. Our study demonstrates that the downregulation of miR-1, miR-133a, and upregulation of miR-21 can be reversed by one single upstream regulator, SRF. These results may help to develop novel therapeutic interventions targeting microRNA biogenesis.

Previous studies have shown that a mitogen activated protein (MAP) kinase (MEK)-independent signaling pathway is required by activated Raf or fibroblast-derived growth factor (FGF) for the differentiation of rat hippocampal neuronal H19-7 cells. We now demonstrate that both Raf and FGF similarly induce prolonged transcription and translation of the immediate early gene pip92 in the absence of activation of the MAP kinases (MAPKs) ERK1 and ERK2. To determine the mechanism by which this occurs and to identify novel Raf-activated signaling pathways, we investigated the induction of the pip92 promoter by both FGF and an estradiol-activated Raf-1–estrogen receptor fusion protein (ΔRaf-1:ER) in H19-7 cells. Deletion analysis of the pip92 promoter indicated that activation by the MAPK-independent pathway occurs primarily within the region containing a serum response element (SRE). Further analysis of the SRE by using a heterologous thymidine kinase promoter showed that both an Ets and CArG...

Summary Several studies have shown that altered gravity con- ditions influence mammalian cell growth and differ- entiation. The molecular mechanisms underlying these effects, however, remain relatively obscure. In this paper we show that microgravity reached in a sounding rocket strongly decreases epidermal growth factor (EGF)-induced expression of the proto- oncogenes c-fos and c-jun, which are both implicated in the regulation of proliferation and differentiation. Decreased activity of the serum response element (SRE), present in the c-fos promoter-enhancer re- gion, is probably responsible for the decrease in EGF-induced c-fos expression. In addition, we show that gravity alterations differentially modulate dis- tinctive signal transduction pathways, indicating that gravity-dependent modulations of mammalian cell proliferation are unlikely to be caused by a non- specific stress response of the cell.

Ms1/STARS is a novel muscle-specific actin-binding protein that specifically modulates the myocardin-related transcription factor (MRTF)-serum response factor (SRF) regulatory axis within striated muscle. This ms1/STARS-dependent regulatory axis is of central importance within the cardiac gene regulatory network and has been implicated in cardiac development and postnatal cardiac function/homeostasis. The dysregulation of ms1/STARS is associated with and causative of pathological cardiac phenotypes, including cardiac hypertrophy and cardiomyopathy. In order to gain an understanding of the mechanisms governing ms1/STARS expression in the heart, we have coupled a comparative genomic in silico analysis with reporter, gain-of-function, and loss-of-function approaches. Through this integrated analysis, we have identified three evolutionarily conserved regions (ECRs), α, SINA, and DINA, that act as cis-regulatory modules and confer differential cardiac cell-specific activity. Two of these...