Ovol1 represses its own transcription by competing with transcription activator c-Myb and by recruiting histone deacetylase activity

Ovol1 belongs to a family of evolutionarily conserved zinc finger proteins that act downstream of key developmental signaling pathways such as Wnt and TGF-β/BMP. It plays important roles in epithelial and germ cell development, particularly by repressing c-Myc and Id2 genes and modulating the balance between proliferation and differentiation of progenitor cells. In this study, we show that Ovol1 negatively regulates its own expression by binding to and repressing the activity of its promoter. We further demonstrate that Ovol1 uses both passive and active repression mechanisms to auto-repress: (1) it antagonizes transcriptional activation of c-Myb, a known positive regulator of proliferation, by competing for DNA binding; (2) it recruits histone deacetylase activity to the promoter via an N-terminal SNAG repressor domain. At Ovol1 cognate sites in the endogenous Ovol1 promoter, c-Myb binding correlates with increased histone acetylation, whereas the expression of Ovol1 correlates with a displacement of c-Myb from the DNA and decreased histone acetylation. Collectively, our data suggest that Ovol1 restricts its own expression by counteracting c-Myb activation and histone acetylation of the Ovol1 promoter

Ovol1 regulates the growth arrest of embryonic epidermal progenitor cells and represses c-myc transcription

Transcriptional control plays a key role in regulating epidermal proliferation and differentiation. Although ample information has been obtained on how epidermal homeostasis is controlled in adult skin, less is known about the control of proliferation/differentiation of epidermal stem/progenitor cells in the developing embryo. Ovol1, encoding a zinc finger protein homologous to Drosophila melanogaster Ovo, is expressed in embryonic epidermal progenitor cells that are transiting from proliferation to terminal differentiation. In this study, we demonstrate a function for Ovol1 in interfollicular epidermal development. In its absence, developing epidermis fails to properly restrict the proliferative potential of progenitor cells, and cultured keratinocytes fail to efficiently undergo growth arrest in response to extrinsic growth-inhibitory signals. We present molecular evidence that c-myc expression is up-regulated in Ovol1-deficient suprabasal cells and that Ovol1 represses c-myc transcription by directly binding to its promoter. Collectively, our findings indicate that Ovol1 is required for proliferation exit of committed epidermal progenitor cells and identify c-myc as an Ovol1 target

(5S,6S,10R)-10-(2,4-Dichloro­phen­yl)-14-[(E)-(2,4-dichloro­phen­yl)methyl­idene]-3,9-diphenyl-12-[(R)-1-phenyl­ethyl]-1,4,7-trioxa-2,8,12-triaza­dispiro­[4.0.4.4]tetra­deca-2,8-diene

The asymmetric unit of the title compound, C41H31Cl4N3O3, contains two independent mol­ecules with almost identical geometries. The piperidine ring adopts a chair conformation in both mol­ecules, and the dihydro­isoxazole rings adopt envelope conformations. The crystal structure is stabilized by C—H⋯N hydrogen bonds and C—H⋯π inter­actions

Opposing effects of Ovol1 and c-Myb on histone H3 acetylation at the endogenous promoter

<p><b>Copyright information:</b></p><p>Taken from "Ovol1 represses its own transcription by competing with transcription activator c-Myb and by recruiting histone deacetylase activity"</p><p></p><p>Nucleic Acids Research 2007;35(5):1687-1697.</p><p>Published online 20 Feb 2007</p><p>PMCID:PMC1865076.</p><p>© 2007 The Author(s).</p> () Results of ChIP assays showing that c-Myb and Ovol1 occupancy correlates with increased and decreased respectively histone H3 acetylation from the basal level. () Results of quantification of ChIP signals. Error bars were calculated from three independent PCR reactions of a single immunoprecipitate, and results are representative of multiple ChIP experiments. ‘*’ indicates statistically significant ( < 0.003) difference from the untransfected sample

Ovol1 binds to its own promoter and binding requires the zinc finger domain

<p><b>Copyright information:</b></p><p>Taken from "Ovol1 represses its own transcription by competing with transcription activator c-Myb and by recruiting histone deacetylase activity"</p><p></p><p>Nucleic Acids Research 2007;35(5):1687-1697.</p><p>Published online 20 Feb 2007</p><p>PMCID:PMC1865076.</p><p>© 2007 The Author(s).</p> () Organization of the promoter showing the positions of the proximal () and distal () sites, with their sequences shown below the stick diagram. The transcription start site is +1. Also indicated are the positions of primer sets 1 and 2 used for ChIP assays (see below). () Results of EMSA assays showing binding of recombinant Ovol1 to CCGTTA-containing oligonucleotides (left) and (right). Left panel: lane 1, free probe, lanes 2–4, with increasing amounts (172–688 nM) of recombinant His–Ovol1. Right panel: lanes 1 and 7, free probe; lanes 2–6, with increasing concentrations, (50–431 nM) of recombinant His–Ovol1; lanes 8 and 9, with 156 nM recombinant His–Ovol1 and with and without anti-Ovol1 antibody, respectively. Arrowhead and arrow represent Ovol1–DNA and Ovol1–DNA–antibody complexes, respectively. () Results of EMSA assays on using recombinant full-length and truncated Ovol1 proteins. Oval and arrowhead represent full-length and C-terminal Ovol1–DNA complexes, respectively. Lower bands (indicated by *) are likely due to incompletely translated protein products. () DNase1 footprint of the promoter. The sequence of the footprinted region (−53 to −35) is indicated on the right. The bracket on the left indicates the position of the low-affinity site which is present within the oligonucleotide . () Densitometer tracing of hydroxyl radical footprint of the promoter, with the footprinted nucleotides underlined and its position indicated