CHIP Represses Myocardin-Induced Smooth Muscle Cell Differentiation via Ubiquitin-Mediated Proteasomal Degradation

Abstract

Myocardin, a coactivator of serum response factor (SRF), plays a critical role in the differentiation of vascular smooth muscle cells (SMCs). However, the molecular mechanisms regulating myocardin stability and activity are not well defined. Here we show that the E3 ligase C terminus of Hsc70-interacting protein (CHIP) represses myocardin-dependent SMC gene expression and transcriptional activity. CHIP interacts with and promotes myocardin ubiquitin-mediated degradation by the proteasome in vivo and in vitro. Furthermore, myocardin ubiquitination by CHIP requires its phosphorylation. Importantly, CHIP overexpression reduces the level of myocardin-dependent SMC contractile gene expression and diminishes arterial contractility ex vivo. These findings for the first time, to our knowledge, demonstrate that CHIP-promoted proteolysis of myocardin plays a key role in the physiological control of SMC phenotype and vessel tone, which may have an important implication for pathophysiological conditions such as atherosclerosis, hypertension, and Alzheimer's disease.Phenotypic modulation of vascular smooth muscle (SM) cells (SMCs) plays a pivotal role in vascular development and remodeling during diseases. It is well established that the SMC phenotype is regulated by a wide range of extracellular cues. In response to vascular injury, vascular SMCs (VSMCs) dedifferentiate to a proliferative phenotype that is required for the various pathological states in atherosclerosis, neointimal hyperplasia, and hypertension (27,28). VSMC differentiation is characterized by expressing the highest levels of SM contractile genes, whereas proliferating SMCs express reduced levels of these genes. Thus, changes in SM contractile gene expression are often used to mark SMC phenotypes. It has been known that most SM contractile genes such as SM ␣-actin, SM myosin heavy chain (SM-MHC), and SM22␣ are controlled by serum response factor (SRF), which binds to a sequence known as a CArG box (18, 28). The specific genes activated by SRF are determined by the intracellular signals, as well as the availability of positive and negative cofactors. One of these factors is the SRF cofactor myocardin, which activates SRF-dependent genes and functions as a "master regulator" of SMC differentiation during development (31,32,36).Myocardin is expressed specifically in SM and cardiac muscle lineages and belongs to a SAP (SAF-A/B, acinus, PIAS) superfamily which has been implicated in cardiovascular development and adaptation of the cardiovascular system to hemodynamic stress (18,28). Myocardin physically interacts with the MADS box transcription factor SRF, activating a subset of genes involved in cardiomyocyte and SMC differentiation in vivo (32). Recent reports from myocardin gain-and loss-offunction studies demonstrate that it is necessary and sufficient to initiate SMC differentiation (32). One study by Huang et al. shows that myocardin conditional mutant mice exhibit markedly diminished expression of SMC contractile proteins in the ductus arteriosus and a gen...

Modulation of Myocardin Function by the Ubiquitin E3 Ligase UBR5

“…It will be very important to assess UBR5 expression or mutation in the smooth muscle cells of atherosclerotic plaque in human patients as this may provide important clinical correlations to these basic and translational observations. Previous studies have demonstrated myocardin expression can be tightly regulated by proteasome-mediated degradation (27,36). For example, the E3 ligase COOH terminus of Hsc70-interacting protein has been found to interact with the COOHterminal transactivation domain of myocardin and repress myocardin-dependent SMC gene expression by promoting degradation of myocardin protein.…”

Role of the Ubiquitin Proteasome System in the Heart

“…118 Additionally, the mind bomb (MIB) family of ubiquitin ligases are essential for Notch signaling by mediating the internalization ubiquitination and degradation of membranous Delta. 109 Conversely, several E3 ligases for Notch have been identified. Fbxw7 targets Notch to the proteasome for degradation in the vasculature, 119 in addition to its well-characterized negative regulation of apoptotic JNK signaling in neurons.…”

“…107 Such an antip53 approach was associated with prevention of myocardial apoptosis and improved LV remodeling. In addition to p53, CHIP also mediates the degradation of myocardin, a key transcription factor of serum response factor, thereby decreasing smooth muscle cell differentiation, 109 and mediates the degradation of FoxO1 (as does Atrogin/MAFbx). Overexpression of CHIP represses FoxO1-mediated transactivation and its proapoptotic function after tumor necrosis factor-α treatment, whereas CHIP knockdown enhances FoxO1-mediated transactivation and its effect on SMC proliferation and survival.…”

STAT3 Protein Regulates Vascular Smooth Muscle Cell Phenotypic Switch by Interaction with Myocardin

“…Although myocardin is considered as a critical component of the contractile "switch" in SMC, the signals that regulate myocardin are only partially understood. Extracellular signals can mediate this molecular switch through alterations in effector pathways, and indeed myocardin-mediated gene activation is regulated by ligands such as TGF␤1 and angiotensin II (16), as well as intracellular signal transduction mediators such as YAP1, glycogen synthase kinase 3 (GSK3), and extracellular signal-regulated kinase (ERK) and the C terminus of Hsc70-interacting protein (ChIP) (17)(18)(19)(20). Although the regulation of myocardin gene expression has been widely explored, there are few studies that address the cellular signal pathways underlying the regulation of myocardin-mediated SMC phenotypic changes.…”