A Non-Canonical Hippo Pathway Represses the Expression of ΔNp63.

The p63 transcription factor, a member of the p53 family, plays an oncogenic role in squamous cell carcinomas, while in breast cancers its expression is often repressed. In the canonical conserved Hippo pathway, known to play a complex role in regulating growth of cancer cells, protein kinases MST1/2 and LATS1/2 act sequentially to phosphorylate and inhibit the YAP/TAZ transcription factors. We found that in MCF10A mammary epithelial cells as well as in squamous and breast cancer cell lines, expression of ΔNp63 RNA and protein is strongly repressed by inhibition of the Hippo pathway protein kinases.

A non-canonical Hippo pathway represses the expression of ΔNp63.

Unlabelled: The p63 transcription factor, a member of the p53 family, plays an oncogenic role in squamous cancers, while in breast cancers its expression is often repressed. In the canonical conserved Hippo pathway, known to play a complex role in regulating growth of cancer cells, the protein kinases MST1/2 and LATS1/2 act sequentially to phosphorylate and inhibit the YAP/TAZ transcription factors. We found that in the MCF10A mammary epithelial cell line as well as in squamous and breast cancer cell lines, expression of ΔNp63 RNA and protein is strongly repressed by inhibition of the Hippo pathway protein kinases in a manner that is independent of p53.

The myocardin-related transcription factor MKL co-regulates the cellular levels of two profilin isoforms.

Megakaryoblastic leukemia (MKL)/serum-response factor (SRF)-mediated gene transcription is a highly conserved mechanism that connects dynamic reorganization of the actin cytoskeleton to regulation of expression of a wide range of genes, including SRF itself and many important structural and regulatory components of the actin cytoskeleton. In this study, we examined the possible role of MKL/SRF in the context of regulation of profilin (Pfn), a major controller of actin dynamics and actin cytoskeletal remodeling in cells. We demonstrated that despite being located on different genomic loci, two major isoforms of Pfn (Pfn1 and Pfn2) are co-regulated by a common mechanism involving the action of MKL that is independent of its SRF-related activity.

Repression of p63 and induction of EMT by mutant Ras in mammary epithelial cells.

The p53-related transcription factor p63 is required for maintenance of epithelial cell differentiation. We found that activated forms of the Harvey Rat Sarcoma Virus GTPase (H-RAS) and phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) oncogenes strongly repress expression of ∆Np63α, the predominant p63 isoform in basal mammary epithelial cells. This regulation occurs at the transcriptional level, and a short region of the ∆Np63 promoter is sufficient for repression induced by H-RasV12.

Filamin A interacts with the coactivator MKL1 to promote the activity of the transcription factor SRF and cell migration.

Megakaryoblastic leukemia 1 (MKL1) is a coactivator of serum response factor (SRF) that promotes the expression of genes associated with cell proliferation, motility, adhesion, and differentiation-processes that also involve dynamic cytoskeletal changes in the cell. MKL1 is inactive when bound to monomeric globular actin (G-actin), but signals that activate the small guanosine triphosphatase RhoA cause actin polymerization and MKL1 dissociation from G-actin. We found a new mechanism of MKL1 activation that is mediated through its binding to filamin A (FLNA), a protein that binds filamentous actin (F-actin).

A novel inhibitory mechanism of MRTF-A/B on the ICAM-1 gene expression in vascular endothelial cells.

The roles of myocardin-related transcription factor A (MRTF-A) and MRTF-B in vascular endothelial cells are not completely understood. Here, we found a novel regulatory mechanism for MRTF-A/B function. MRTF-A/B tend to accumulate in the nucleus in arterial endothelial cells in vivo and human aortic endothelial cells (HAoECs) in vitro.

Iron deficiency upregulates Egr1 expression.

Iron-deficient anemia is a prevalent disease among humans. We searched for genes regulated by iron deficiency and its regulated mechanism. cDNA microarrays were performed using Hepa1c1c7 cells treated with 100 μM desferrioxamine (DFO), an iron chelator.

Pathway Regulation of p63, a Director of Epithelial Cell Fate.

The p53-related gene p63 is required for epithelial cell establishment and its expression is often altered in tumor cells. Great strides have been made in understanding the pathways and mechanisms that regulate p63 levels, such as the Wnt, Hedgehog, Notch, and EGFR pathways. We discuss here the multiple signaling pathways that control p63 expression as well as transcription factors and post-transcriptional mechanisms that regulate p63 levels.

Serum regulation of Id1 expression by a BMP pathway and BMP responsive element.

Immediate early genes (IEGs) are expressed upon re-entry of quiescent cells into the cell cycle following serum stimulation. These genes are involved in growth control and differentiation and hence their expression is tightly controlled. Many IEGs are regulated through Serum Response Elements (SREs) in their promoters, which bind Serum Response Factor (SRF).

Depletion of the transcriptional coactivators megakaryoblastic leukaemia 1 and 2 abolishes hepatocellular carcinoma xenograft growth by inducing oncogene-induced senescence.

Megakaryoblastic leukaemia 1 and 2 (MKL1/2) are coactivators of the transcription factor serum response factor (SRF). Here, we provide evidence that depletion of MKL1 and 2 abolishes hepatocellular carcinoma (HCC) xenograft growth. Loss of the tumour suppressor deleted in liver cancer 1 (DLC1) and the subsequent activation of RhoA were prerequisites for MKL1/2 knockdown-mediated growth arrest.

Fra-1 regulation of Matrix Metallopeptidase-1 (MMP-1) in metastatic variants of MDA-MB-231 breast cancer cells.

Matrix Metallopeptidase 1 (MMP-1) expression has repeatedly been correlated to tumorigenesis and metastasis.  Yet, MMP-1 regulation in a metastatic context remains largely unknown.  Here we confirm differential MMP-1 expression in mammary carcinoma cells with varied metastatic potentials.

An IFN-γ-stimulated ATF6-C/EBP-β-signaling pathway critical for the expression of Death Associated Protein Kinase 1 and induction of autophagy.

The IFN family of cytokines operates a frontline defense against pathogens and neoplastic cells in vivo by controlling the expression of several genes. The death-associated protein kinase 1 (DAPK1), an IFN-γ-induced enzyme, controls cell cycle, apoptosis, autophagy, and tumor metastasis, and its expression is frequently down-regulated in a number of human tumors. Although the biochemical action of DAPK1 is well understood, mechanisms that regulate its expression are unclear.

Activation and repression of cellular immediate early genes by serum response factor cofactors.

The induction of expression of many cellular immediate early genes (IEG) involves the transcription factor serum response factor (SRF). Two families of SRF coactivators have also been implicated in IEG induction, the ternary complex factors (TCFs), ELK1, Sap1, and Net, and the myocardin-related factors, MKL1 and MKL2. We found that serum induction of some SRF target genes is preferentially regulated by MKL1/2, whereas others are redundantly activated by both TCFs and MKL1/2.

Serum-induced phosphorylation of the serum response factor coactivator MKL1 by the extracellular signal-regulated kinase 1/2 pathway inhibits its nuclear localization.

Megakaryoblastic leukemia 1 (MKL1) is a myocardin-related coactivator of the serum response factor (SRF) transcription factor, which has an integral role in differentiation, migration, and proliferation. Serum induces RhoA-dependent translocation of MKL1 from the cytoplasm to the nucleus and also causes a rapid increase in MKL1 phosphorylation. We have mapped a serum-inducible phosphorylation site and found, surprisingly, that its mutation causes constitutive localization to the nucleus, suggesting that phosphorylation of MKL1 inhibits its serum-induced nuclear localization.

ER stress signaling by regulated proteolysis of ATF6.

ATF6 is an endoplasmic reticulum (ER) membrane-anchored transcription factor activated by intramembrane proteolysis in the ER stress response. Upon ER stress, ATF6 is transported from the ER to the Golgi to be processed by site-1 and site-2 proteases. The trafficking is controlled by the ER chaperone BiP/GRP78.

Stable binding of ATF6 to BiP in the endoplasmic reticulum stress response.

Endoplasmic reticulum (ER) stress-induced activation of ATF6, an ER membrane-bound transcription factor, requires a dissociation step from its inhibitory regulator, BiP. It has been generally postulated that dissociation of the BiP-ATF6 complex is a result of the competitive binding of misfolded proteins generated during ER stress. Here we present evidence against this model and for an active regulatory mechanism for dissociation of the complex.

Myocardin/MKL family of SRF coactivators: key regulators of immediate early and muscle specific gene expression.

Myocardin, megakaryoblastic leukemia-1 (MKL1), and MKL2 belong to a newly defined family of transcriptional coactivators. All three family members bind to serum response factor (SRF) and strongly activate transcription from promoters with SRF binding sites. SRF is required for the serum induction of immediate early genes such as c-fos and for the expression of many muscle specific genes.

Expression profiling of serum inducible genes identifies a subset of SRF target genes that are MKL dependent.

Background: Serum Response Factor (SRF) is a transcription factor that is required for the expression of many genes including immediate early genes, cytoskeletal genes, and muscle-specific genes. SRF is activated in response to extra-cellular signals by its association with a diverse set of co-activators in different cell types. In the case of the ubiquitously expressed immediate early genes, the two sets of SRF binding proteins that regulate its activity are the TCF family of proteins that include Elk1, SAP1 and SAP2 and the myocardin-related MKL family of proteins that include MKL1 and MKL2 (also known as MAL, MRTF-A and -B and BSAC).

Dependence of site-2 protease cleavage of ATF6 on prior site-1 protease digestion is determined by the size of the luminal domain of ATF6.

ATF6 is an endoplasmic reticulum (ER) membrane-anchored transcription factor activated by regulated intramembrane proteolysis in the ER stress response. The release of the cytosolic transcription factor domain of ATF6 requires the sequential processing by the Golgi site-1 and site-2 proteases (S1P and S2P). It has been unclear why S2P proteolysis relies on previous site-1 cleavage.

Megakaryoblastic leukemia-1/2, a transcriptional co-activator of serum response factor, is required for skeletal myogenic differentiation.

Serum response factor (SRF) is required for the expression of a wide variety of muscle-specific genes that are expressed upon differentiation and is thus required for both striated and smooth muscle differentiation in addition to its role in regulating growth factor-inducible genes. A heart and smooth muscle-specific SRF co-activator, myocardin, has been shown to be required for cardiac development and smooth muscle differentiation. However, no such co-factors of SRF have been identified in the skeletal myogenic differentiation program.

Megakaryoblastic leukemia 1, a potent transcriptional coactivator for serum response factor (SRF), is required for serum induction of SRF target genes.

Megakaryoblastic leukemia 1 (MKL1) is a myocardin-related transcription factor that we found strongly activated serum response element (SRE)-dependent reporter genes through its direct binding to serum response factor (SRF). The c-fos SRE is regulated by mitogen-activated protein kinase phosphorylation of ternary complex factor (TCF) but is also regulated by a RhoA-dependent pathway. The mechanism of this pathway is unclear.

ATF1 phosphorylation by the ERK MAPK pathway is required for epidermal growth factor-induced c-jun expression.

Epidermal growth factor induction of c-jun expression requires ATF1 and MEF2 sites in the c-jun promoter. We find that activation of the c-jun promoter through the ATF1 site requires phosphorylation of ATF1 at serine 63. A serine 63 to alanine mutation of ATF1 acts to block epidermal growth factor (EGF) induction of a transfected c-jun gene.

ER stress regulation of ATF6 localization by dissociation of BiP/GRP78 binding and unmasking of Golgi localization signals.

ATF6 is an endoplasmic reticulum (ER) stress-regulated transmembrane transcription factor that activates the transcription of ER molecular chaperones. Upon ER stress, ATF6 translocates from the ER to the Golgi where it is processed to its active form. We have found that the ER chaperone BiP/GRP78 binds ATF6 and dissociates in response to ER stress.

Composition and function of AP-1 transcription complexes during muscle cell differentiation.

The role of activating protein-1 (AP-1) in muscle cells is currently equivocal. While some studies propose that AP-1 is inhibitory for myogenesis, others implicate a positive role in this process. We tested whether this variation may be due to different properties of the AP-1 subunit composition in differentiating cells.