HDAC5 is required for maintenance of pericentric heterochromatin and controls cell cycle progression of human cancer cells

Poster (2012, February 04)

The angiogenesis suppressor gene AKAP12 is under the epigenetic control of HDAC7 in endothelial cells.

in Angiogenesis (2012)

Histone deacetylases (HDACs) are a family of 18 enzymes that deacetylate lysine residues of both histone and nonhistone proteins and to a large extent govern the process of angiogenesis. Previous studies ... [more ▼]

Histone deacetylases (HDACs) are a family of 18 enzymes that deacetylate lysine residues of both histone and nonhistone proteins and to a large extent govern the process of angiogenesis. Previous studies have shown that specific inhibition of HDAC7 blocks angiogenesis both in vitro and in vivo. However, the underlying molecular mechanisms are not fully understood and hence preclude any meaningful development of suitable therapeutic modalities. The goal of the present study was to further the understanding of HDAC7 epigenetic control of angiogenesis in human endothelial cells using the proteomic approach. The underlying problem was approached through siRNA-mediated gene-expression silencing of HDAC7 in human umbilical vein endothelial cells (HUVECs). To this end, HUVEC proteins were extracted and proteomically analyzed. The emphasis was placed on up-regulated proteins, as these may represent potential direct epigenetic targets of HDAC7. Among several proteins, A-kinase anchor protein 12 (AKAP12) was the most reproducibly up-regulated protein following HDAC7 depletion. This overexpression of AKAP12 was responsible for the inhibition of migration and tube formation in HDAC7-depleted HUVEC. Mechanistically, H3 histones associated with AKAP12 promoter were acetylated following the removal of HDAC7, leading to an increase in its mRNA and protein levels. AKAP12 is responsible for protein kinase C mediated phosphorylation of signal transducer and activator of transcription 3 (STAT3). Phosphorylated STAT3 increasingly binds to the chromatin and AKAP12 promoter and is necessary for maintaining the elevated levels of AKAP12 following HDAC7 knockdown. We demonstrated for the first time that AKAP12 tumor/angiogenesis suppressor gene is an epigenetic target of HDAC7, whose elevated levels lead to a negative regulation of HUVEC migration and inhibit formation of tube-like structures. [less ▲]

Histone Deacetylase 7 Regulates Cell Survival and TCR Signaling in CD4/CD8 Double-Positive Thymocytes.

in Journal of Immunology (2011), 186(8), 4782-93

CD4/CD8 double-positive thymocytes express the transcriptional repressor histone deacetylase (HDAC)7, a class IIa HDAC that is exported from the cell nucleus after TCR engagement. Through signal-dependent ... [more ▼]

CD4/CD8 double-positive thymocytes express the transcriptional repressor histone deacetylase (HDAC)7, a class IIa HDAC that is exported from the cell nucleus after TCR engagement. Through signal-dependent nuclear export, class IIa HDACs such as HDAC7 mediate signal-dependent changes in gene expression that are important to developmental fate decisions in multiple tissues. We report that HDAC7 is exported from the cell nucleus during positive selection in mouse thymocytes and that it regulates genes mediating the coupling between TCR engagement and downstream events that determine cell survival. Thymocytes lacking HDAC7 are inefficiently positively selected due to a severely shortened lifespan and exhibit a truncated repertoire of TCR Jalpha segments. The expression of multiple important mediators and modulators of the response to TCR engagement is altered in HDAC7-deficient thymocytes, resulting in increased tonic MAPK activity that contributes to the observed loss of viability. Remarkably, the activity of protein kinase D, the kinase that mediates nuclear export of HDAC7 in response to TCR signaling, is also increased in HDAC7-deficient thymocytes, suggesting that HDAC7 nuclear export governs a self-sustaining autoexcitatory loop. These experiments add to the understanding of the life/death decision in thymic T cell development, define a novel function for class IIa HDACs, and point to a novel feed-forward mechanism whereby these molecules regulate their own state and mediate stable developmental transitions. [less ▲]

Dentin Matrix Protein 1 induces membrane expression of VE-cadherin on endothelial cells and inhibits VEGF-induced angiogenesis by blocking VEGFR-2 phosphorylation.

in Blood (2011), 117(8), 2515-26

Dentin matrix protein 1 (DMP1) is a member of the Small Integrin-Binding LIgand N-linked Glycoproteins (SIBLINGs) family, a group of proteins initially described as mineralized extracellular matrices ... [more ▼]

Dentin matrix protein 1 (DMP1) is a member of the Small Integrin-Binding LIgand N-linked Glycoproteins (SIBLINGs) family, a group of proteins initially described as mineralized extracellular matrices components. More recently, SIBLINGs have been implicated in several key steps of cancer progression, including angiogenesis. Although pro-angiogenic activities have been demonstrated for two SIBLINGs, the role of DMP1 in angiogenesis has not been addressed yet. We demonstrated that this extracellular matrix protein induced the expression of VE-cadherin, a key regulator of intercellular junctions and contact inhibition of growth of endothelial cells that is also known to modulate VEGFR-2 activity, the major high affinity receptor for VEGF. DMP1 induced VE-cadherin and p27(Kip1) expression followed by cell cycle arrest in human umbilical vein endothelial cells (HUVEC) in a CD44-dependent manner. VEGF-induced proliferation, migration and tubulogenesis responses were specifically blocked upon DMP1 pre-treatment of HUVEC. Indeed, subsequently to VE-cadherin induction, DMP1 inhibited VEGFR-2 phosphorylation and Src-mediated signaling. However, DMP1 did not interfere with bFGF-induced angiogenesis. In vivo, DMP1 significantly reduced laser-induced choroidal neovascularization lesions and tumor-associated angiogenesis. These data enable us to put DMP1 on the angiogenic chessboard for the first time and to identify this protein as a new specific inhibitor of VEGF-induced angiogenesis. [less ▲]

HDAC4 represses p21(WAF1/Cip1) expression in human cancer cells through a Sp1-dependent, p53-independent mechanism.

in Oncogene (2009), 28(2), 243-56

Cancer cells have complex, unique characteristics that distinguish them from normal cells, such as increased growth rates and evasion of anti-proliferative signals. Global inhibition of class I and II ... [more ▼]

Cancer cells have complex, unique characteristics that distinguish them from normal cells, such as increased growth rates and evasion of anti-proliferative signals. Global inhibition of class I and II histone deacetylases (HDACs) stops cancer cell proliferation in vitro and has proven effective against cancer in clinical trials, at least in part, through transcriptional reactivation of the p21(WAF1/Cip1)gene. The HDACs that regulate p21(WAF1/Cip1) are not fully identified. Using small interfering RNAs, we found that HDAC4 participates in the repression of p21(WAF1/Cip1) through Sp1/Sp3-, but not p53-binding sites. HDAC4 interacts with Sp1, binds and reduces histone H3 acetylation at the Sp1/Sp3 binding site-rich p21(WAF1/Cip1) proximal promoter, suggesting a key role for Sp1 in HDAC4-mediated repression of p21(WAF1/Cip1). Induction of p21(WAF1/Cip1) mediated by silencing of HDAC4 arrested cancer cell growth in vitro and inhibited tumor growth in an in vivo human glioblastoma model. Thus, HDAC4 could be a useful target for new anti-cancer therapies based on selective inhibition of specific HDACs. [less ▲]

Histone deacetylase 7 silencing alters endothelial cell migration, a key step in angiogenesis

in Circulation Research (2007), 101(12), 1237-1246

Global inhibition of class I and II histone deacetylases (HDACs) impairs angiogenesis. Herein, we have undertaken the identification of the specific HDAC(s) with activity that is necessary for the ... [more ▼]

Global inhibition of class I and II histone deacetylases (HDACs) impairs angiogenesis. Herein, we have undertaken the identification of the specific HDAC(s) with activity that is necessary for the development of blood vessels. Using small interfering RNAs, we observed that HDAC7 silencing in endothelial cells altered their morphology, their migration, and their capacity to form capillary tube-like structures in vitro but did not affect cell adhesion, proliferation, or apoptosis. Among several factors known to be involved in angiogenesis, platelet-derived growth factor-B (PDGF-B) and its receptor (PDGFR-beta) were the most upregulated genes following HDAC7 silencing. We demonstrated that their increased expression induced by HDAC7 silencing was partially responsible for the inhibition of endothelial cell migration. In addition, we have also shown that treatment of endothelial cells with phorbol 12-myristate 13-acetate resulted in the exportation of HDAC7 out of the nucleus through a protein kinase C/protein kinase D activation pathway and induced, similarly to HDAC7 silencing, an increase in PDGF-B expression, as well as a partial inhibition of endothelial cell migration. Collectively, these data identified HDAC7 as a key modulator of endothelial cell migration and hence angiogenesis, at least in part, by regulating PDGF-B/PDGFR-beta gene expression. Because angiogenesis is required for tumor progression, HDAC7 may represent a rational target for therapeutic intervention against cancer. [less ▲]

Role for casein kinase 2 in the regulation of HIF-1 activity.

in International Journal of Cancer = Journal International du Cancer (2005), 117(5), 764-74

Hypoxia-inducible factor-1 (HIF-1) is a heterodimeric transcription factor that plays a major role in cellular adaptation to hypoxia. The mechanisms regulating HIF-1 activity occurs at multiple levels in ... [more ▼]

Hypoxia-inducible factor-1 (HIF-1) is a heterodimeric transcription factor that plays a major role in cellular adaptation to hypoxia. The mechanisms regulating HIF-1 activity occurs at multiple levels in vivo. The HIF-1alpha subunit is highly sensible to oxygen and is rapidly degraded by the proteasome 26S in normoxia. Activation in hypoxia occurs through a multistep process including inhibition of HIF-1alpha degradation, but also increase in the transactivation activity of HIF-1. Several data indicate that phosphorylation could play a role in this regulation. In this report, we investigated the role of casein kinase 2 (CK2), an ubiquitous serine/threonine kinase, in the regulation of HIF-1 activity. Hypoxia was capable of increasing the expression of the beta subunit of CK2, of inducing a relocalization of this subunit at the plasma membrane, of inducing nuclear translocation of the alpha subunit and of increasing CK2 activity. Three inhibitors of this kinase, DRB (5,6-dichloro-1-beta-D-ribofuranosyl-benzimidazole), TBB (4,5,6,7-tetrabromotriazole) and apigenin, as well as overexpression of a partial dominant negative mutant of CK2alpha, were shown to inhibit HIF-1 activity as measured by a reporter assay and through hypoxia-induced VEGF and aldolase expression. This does not occur at the stabilization process since they did not affect HIF-1alpha protein level. DNA-binding activity was also not inhibited. We conclude that CK2 is an important regulator of HIF-1 transcriptional activity but the mechanism of this regulation remains to be determined. Since HIF-1 plays a major role in tumor angiogenesis and since CK2 has been described to be overexpressed in tumor cells, this new pathway of regulation can be one more way for tumor cells to survive. [less ▲]

Regulation of hypoxia-inducible factor-1alpha protein level during hypoxic conditions by the phosphatidylinositol 3-kinase/Akt/glycogen synthase kinase 3beta pathway in HepG2 cells.

in Journal of Biological Chemistry (2003), 278(33), 31277-85

Hypoxia initiates an intracellular signaling pathway leading to the activation of the transcription factor hypoxia-inducible factor-1 (HIF-1). HIF-1 activity is regulated through different mechanisms ... [more ▼]

Hypoxia initiates an intracellular signaling pathway leading to the activation of the transcription factor hypoxia-inducible factor-1 (HIF-1). HIF-1 activity is regulated through different mechanisms involving stabilization of HIF-1alpha, phosphorylations, modifications of redox conditions, and interactions with coactivators. However, it appears that some of these steps can be cell type-specific. Among them, the involvement of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in the regulation of HIF-1 by hypoxia remains controversial. Here, we investigated the activation state of PI3K/Akt/glycogen synthase kinase 3beta (GSK3beta) in HepG2 cells. Increasing incubation times in hypoxia dramatically decreased both the phosphorylation of Akt and the inhibiting phosphorylation of GSK3beta. The PI3K/Akt pathway was necessary for HIF-1alpha stabilization early during hypoxia. Indeed, its inhibition was sufficient to decrease HIF-1alpha protein level after 5-h incubation in hypoxia. However, longer exposure (16 h) in hypoxia resulted in a decreased HIF-1alpha protein level compared with early exposure (5 h). At that time, Akt was no longer present or active, which resulted in a decrease in the inhibiting phosphorylation of GSK3beta on Ser-9 and hence in an increased GSK3beta activity. GSK3 inhibition reverted the effect of prolonged hypoxia on HIF-1alpha protein level; more stabilized HIF-1alpha was observed as well as increased HIF-1 transcriptional activity. Thus, a prolonged hypoxia activates GSK3beta, which results in decreased HIF-1alpha accumulation. In conclusion, hypoxia induced a biphasic effect on HIF-1alpha stabilization with accumulation in early hypoxia, which depends on an active PI3K/Akt pathway and an inactive GSK3beta, whereas prolonged hypoxia results in the inactivation of Akt and activation of GSK3beta, which then down-regulates the HIF-1 activity through down-regulation of HIF-1alpha accumulation. [less ▲]

Is HIF-1alpha a pro- or an anti-apoptotic protein?

in Biochemical Pharmacology (2002), 64(5-6), 889-92

Hypoxia-inducible factor-1 (HIF-1) is the major transcription factor specifically activated by hypoxia. It induces the expression of different genes whose products play an adaptive role for hypoxic cells ... [more ▼]

Hypoxia-inducible factor-1 (HIF-1) is the major transcription factor specifically activated by hypoxia. It induces the expression of different genes whose products play an adaptive role for hypoxic cells and tissues. Besides these protective responses, HIF-1 and/or hypoxia have also been shown to be either anti-apoptotic or pro-apoptotic, according to the cell type and experimental conditions. More severe or prolonged hypoxia rather induces apoptosis that is, at least in part, initiated by the direct association of HIF-1alpha and p53 and p53-induced gene expression. On the other hand, HIF-1alpha dimerized with ARNT, as an active transcription factor, can protect cells from apoptosis induced by several conditions. This review is aimed to describe the different mechanisms that account for these opposite effects of HIF-1alpha. [less ▲]

ERK and calcium in activation of HIF-1.

in Annals of the New York Academy of Sciences (2002), 973

HIF-1 (hypoxia-inducible factor-1) is the main transcription factor responsible for increased gene expression in hypoxia. The oxygen-dependent regulation of HIF-1 activity occurs at multiple levels in ... [more ▼]

HIF-1 (hypoxia-inducible factor-1) is the main transcription factor responsible for increased gene expression in hypoxia. The oxygen-dependent regulation of HIF-1 activity occurs at multiple levels in vivo. The mechanisms regulating HIF-1alpha protein expression have been most extensively analyzed, but the ones modulating HIF-1 transcriptional activity remain unclear. Changes in the phosphorylation and/or redox status of HIF-1alpha certainly play a role. Here, we show that ionomycin could activate HIF-1 transcriptional activity in a way that is additive to the effect of hypoxia without affecting HIF-1alpha protein level and HIF-1 DNA binding capacity. In addition, a calmodulin dominant-negative mutant as well as BAPTA, an intracellular calcium chelator, inhibited the hypoxia-induced HIF-1 activation. These results indicate that elevated calcium in hypoxia could participate in HIF-1 activation. PD98059, an inhibitor of the ERK pathway, but not KN-93, an inhibitor of calmodulin kinases II and IV, also blocked HIF-1 activation by hypoxia and by ionomycin. Altogether, these results suggest that calcium and calmodulin would act upstream of ERK in the hypoxia signal transduction pathway leading to enhanced HIF-1 transcriptional activity. [less ▲]