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Franck Dequiedt

Publications and source records attributed to Franck Dequiedt.

12 recordsLinked to original sources
PubMed2006

Human T-cell leukemia virus type-1 Tax oncoprotein regulates G-protein signaling.

Human T-cell leukemia virus type-1 (HTLV-1) is associated with adult T-cell leukemia (ATL) and neurological syndromes. HTLV-1 encodes the oncoprotein Tax-1, which modulates viral and cellular gene expression leading to T-cell transformation. Guanine nucleotide-binding proteins (G proteins) and G protein-coupled receptors (GPCRs) constitute the largest family of membrane proteins known and are involved in the regulation of most biological functions. Here, we report an interaction between HTLV-1 Tax oncoprotein and the G-protein beta subunit. Interestingly, though the G-protein beta subunit inhibits Tax-mediated viral transcription, Tax-1 perturbs G-protein beta subcellular localization. Functional evidence for these observations was obtained using conditional Tax-1-expressing transformed T-lymphocytes, where Tax expression correlated with activation of the SDF-1/CXCR4 axis. Our data indicated that HTLV-1 developed a strategy based on the activation of the SDF-1/CXCR4 axis in the infected cell; this could have tremendous implications for new therapeutic strategies.

Cells, Cultured↗
PubMed2006

New role for hPar-1 kinases EMK and C-TAK1 in regulating localization and activity of class IIa histone deacetylases.

Class IIa histone deacetylases (HDACs) are found both in the cytoplasm and in the nucleus where they repress genes involved in several major developmental programs. In response to specific signals, the repressive activity of class IIa HDACs is neutralized through their phosphorylation on multiple N-terminal serine residues and 14-3-3-mediated nuclear exclusion. Here, we demonstrate that class IIa HDACs are subjected to signal-independent nuclear export that relies on their constitutive phosphorylation. We identify EMK and C-TAK1, two members of the microtubule affinity-regulating kinase (MARK)/Par-1 family, as regulators of this process. We further show that EMK and C-TAK1 phosphorylate class IIa HDACs on one of their multiple 14-3-3 binding sites and alter their subcellular localization and repressive function. Using HDAC7 as a paradigm, we extend these findings by demonstrating that signal-independent phosphorylation of the most N-terminal serine residue by the MARK/Par-1 kinases, i.e., Ser155, is a prerequisite for the phosphorylation of the nearby 14-3-3 site, Ser181. We propose that this multisite hierarchical phosphorylation by a variety of kinases allows for sophisticated regulation of class IIa HDACs function.

14-3-3 Proteins↗
PubMed2005

MAPK pathway contributes to density- and hypoxia-induced expression of the tumor-associated carbonic anhydrase IX.

Transcription of the CA9 gene coding for a tumor-associated carbonic anhydrase IX (CA IX) isoform is regulated by hypoxia via the hypoxia-inducible factor 1 (HIF-1) and by high cell density via the phosphatidylinositol-3-kinase (PI3K) pathway. We examined the role of the mitogen-activated protein kinase (MAPK) pathway in the control of CA9 gene expression. Inhibition of MAPK signaling by U0126 in HeLa cells led to reduced activity of the PR1-HRE-luc CA9 promoter construct and decreased CA IX protein levels in dense culture as well as in hypoxia. Similar reduction was obtained by expression of a dominant-negative ERK1 mutant and was also observed in U0126-treated HIF-1alpha-deficient Ka13 cells. Simultaneous treatment with the MAPK and PI3K inhibitors U0126 and LY 294002 had stronger effect than individual inhibition of these pathways. Taken together, our results suggest that besides the PI3K pathway, the MAPK cascade is involved in the regulation of CA9 gene expression under both hypoxia and high cell density.

Antigens, Neoplasm↗
PubMed2005

Phosphorylation of histone deacetylase 7 by protein kinase D mediates T cell receptor-induced Nur77 expression and apoptosis.

The molecular basis of thymocyte negative selection, a crucial mechanism in establishing central tolerance, is not yet resolved. Histone deacetylases (HDACs) have emerged as key transcriptional regulators in several major developmental programs. Recently, we showed that the class IIa member, HDAC7, regulates negative selection by repressing expression of Nur77, an orphan nuclear receptor involved in antigen-induced apoptosis of thymocytes. Engagement of the T cell receptor (TCR) alleviates this repression through phosphorylation-dependent nuclear exclusion of HDAC7. However, the identity of the TCR-activated kinase that phosphorylates and inactivates HDAC7 was still unknown. Here, we demonstrate that TCR-induced nuclear export of HDAC7 and Nur77 expression is mediated by activation of protein kinase D (PKD). Indeed, active PKD stimulates HDAC7 nuclear export and Nur77 expression. In contrast, inhibition of PKD prevents TCR-mediated nuclear exclusion of HDAC7 and associated Nur77 activation. Furthermore, we show that HDAC7 is an interaction partner and a substrate for PKD. We identify four serine residues in the NH(2) terminus of HDAC7 as targets for PKD. More importantly, a mutant of HDAC7 specifically deficient in phosphorylation by PKD, inhibits TCR-mediated apoptosis of T cell hybridomas. These findings indicate that PKD is likely to play a key role in the signaling pathways controlling negative selection.

Active Transport, Cell Nucleus↗
PubMed2005

Effect of inhibitors of histone deacetylase on the induction of cell differentiation in murine and human erythroleukemia cell lines.

Histone deacetylase (HDAC) inhibitors are a novel class of promising anti-cancer agents. Little information is available on the capacity of structurally different HDAC inhibitors to induce terminal cell differentiation in different cell types in relation to enzyme inhibition and subtype selectivity. Consequently, the aim of this study was to provide a comprehensive comparison of these effects. New biarylalanine inhibitors of HDAC were synthesized and compared to a series of standard inhibitors from different laboratories. Chromatographically purified rat liver and immunoprecipitated FLAG-tagged recombinant human HDACs were used as sources of HDAC activity. Enzyme inhibition was studied using a fluorescent substrate and its conversion was monitored by high-performance liquid chromatography. The ability to induce cell differentiation was compared in murine (Friend DS-19) and human (K562) erythroleukemic cell lines, and was quantified by benzidine staining. Inhibition of cell proliferation was evaluated by cell counting. All HDAC inhibitors were identified as potent inhibitors of erythroleukemic cell proliferation. However, we observed a complex pattern of differentiation induction: structurally similar inhibitors resulted in disparate activity profiles, whereas similar profiles were detected within distinct structural classes. Among the newly synthesized biarylalanine compounds, a 3'-methoxy derivative was identified as a very effective inducer of terminal cell differentiation. We conclude that investigation of subtype selectivity of selected HDAC inhibitors does not provide a clear link between selectivity and the observed cellular activity profile. The predictive value of in vitro HDAC inhibition assays for identifying anti-proliferative compounds has been emphasized.

Aminobiphenyl Compounds↗
PubMed2004

Subtype selective substrates for histone deacetylases.

To probe the steric requirements for deacylation, we synthesized lysine-derived small molecule substrates and examined structure-reactivity relationships with various histone deacetylases. Rat liver, human HeLa, and human recombinant class I and II histone deacetylases (HDACs) as well as human recombinant NAD(+)-dependent SIRT1 (class III enzyme) were used in these studies. A benzyloxycarbonyl substituent on the alpha-amino group yielded the highest conversion rates. Replacing the epsilon-acetyl group with larger lipophilic acyl substituents led to a pronounced decrease in conversion by class I and II enzymes; the class III enzyme displayed a greater tolerance. Incubations with recombinant FLAG-tagged human HDACs 1, 3, and 6 showed a distinct subtype selectivity among small molecule substrates. The subtype selectivity of HDAC inhibitors could be predicted with these substrates and an easily obtainable mixture of HDAC subtypes.

Animals↗
PubMed2003

Nonisotopic substrate for assaying both human zinc and NAD+-dependent histone deacetylases.

Histone deacetylases (HDACs) are involved in the regulation of transcription and their inhibitors are a promising class of new anticancer drugs. We have previously reported Boc(Ac)Lys-AMC, also termed MAL, as a fluorescent substrate for HDACs. Now we present a modification of MAL called Z-MAL that is characterized by an increased rate of conversion by histone deacetylases of classes I and II and the recently discovered sirtuins (histone deacetylases class III). MAL and Z-MAL are the first nonradioactive substrates for class III enzymes. The new substrate Z-MAL allows for shorter assay times in inhibitor screening and is applicable to diverse sources of deacetylase activity even with completely different catalytic mechanisms. Interestingly, MAL shows some relative preference toward class II, indicating that subtype selectivity in small-molecule HDAC substrates might be obtained.

Animals↗
PubMed2003

Class II histone deacetylases: versatile regulators.

Histone acetylation and deacetylation play essential roles in modifying chromatin structure and regulating gene expression in eukaryotes. Histone deacetylases (HDACs) catalyze the deacetylation of lysine residues in the histone N-terminal tails and are found in large multiprotein complexes with transcriptional co-repressors. Human HDACs are grouped into three classes based on their similarity to known yeast factors: class I HDACs are similar to the yeast transcriptional repressor yRPD3, class II HDACs to yHDA1 and class III HDACs to ySIR2. In this review, we focus on the biology of class II HDACs. These newly discovered enzymes have been implicated as global regulators of gene expression during cell differentiation and development. We discuss their emerging biological functions and the molecular mechanisms by which they are regulated.

Animals↗
PubMed2003

HDAC7, a thymus-specific class II histone deacetylase, regulates Nur77 transcription and TCR-mediated apoptosis.

We report that HDAC7, a class II histone deacetylase, is highly expressed in CD4(+)CD8(+) double-positive thymocytes. HDAC7 inhibits the expression of Nur77, an orphan receptor involved in apoptosis and negative selection, via the transcription factor MEF2D. HDAC7 is exported from the nucleus during T cell receptor activation, leading to Nur77 expression. A triple HDAC7 mutant (S155A, S318A, S448A) is not exported from the nucleus in response to TCR activation and suppresses TCR-mediated apoptosis. Conversely, a fusion of HDAC7 to the transcriptional activator VP16 activates Nur77 expression. Inhibition of HDAC7 expression by RNA interference causes increased apoptosis in response to TCR activation. These observations define HDAC7 as a regulator of Nur77 and apoptosis in developing thymocytes.

Acetylation↗
PubMed2003

Transcriptional suppression of in vitro-integrated human immunodeficiency virus type 1 does not correlate with proviral DNA methylation.

Persistence of human immunodeficiency virus type 1 (HIV-1) constitutes a major obstacle in the control of HIV-1 infection. Here we investigated whether the CpG methylation of the HIV-1 promoter can directly influence the expression of the HIV-1 genome and thereby contribute to the persistence and latency of HIV-1. The levels of CpG methylation in the promoter of HIV-1 were studied after bisulfite-induced modification of DNA in five Jurkat clonal cell lines transduced by an HIV-1 long terminal repeat (LTR)-driven retroviral vector and expressing enhanced green fluorescent protein (GFP) and in primary resting memory T cells challenged with HIV-1 or with an HIV-1-derived retroviral vector. Basal HIV-1 promoter activities were low or undetectable in three tested HIV-1 LTR-GFP clones, one of which encoded the Tat protein, and they reached medium or high levels in two other clones. The CpG dinucleotide that occurred in a latently infected clonal cell line 240 nucleotides upstream of the transcription start remained methylated after reactivation of HIV-1 transcription with 10 nM phorbol-12-myristate-13-acetate. In two clones showing a medium promoter activity and in resting memory T cells, the HIV-1 LTR was generally not methylated. Our results show that the methylation profiles of the HIV-1 LTR, including those present in latently infected cells, are low and do not correlate with the transcriptional activity. We suggest that, in a noncloned cellular population in which the HIV-1 proviruses are randomly integrated in the human genome, HIV-1 latency is imperfectly controlled by CpG methylation and is inherently accompanied by residual replication.

Base Sequence↗
PubMed2002

Enzymatic activity associated with class II HDACs is dependent on a multiprotein complex containing HDAC3 and SMRT/N-CoR.

Histone deacetylases (HDACs) play a key role in regulating eukaryotic gene expression. The HDAC domain, homologous to the yeast repressors RPD3 and HDA1, is considered necessary and sufficient for enzymatic activity. Here, we show that the catalytic domain of HDAC4 interacts with HDAC3 via the transcriptional corepressor N-CoR/SMRT. All experimental conditions leading to the suppression of HDAC4 binding to SMRT/N-CoR and to HDAC3 result in the loss of enzymatic activity associated with HDAC4. In vitro reconstitution experiments indicate that HDAC4 and other class II HDACs are inactive in the context of the SMRT/N-CoR-HDAC3 complex and do not contribute to its enzymatic activity. These observations indicate that class II HDACs regulate transcription by bridging the enzymatically active SMRT/N-CoR-HDAC3 complex and select transcription factors independently of any intrinsic HDAC activity.

Amino Acid Sequence↗