RESUMO
The myeolomonocytic cell line U937 differentiates into macrophages in response to a variety of agents. Several genes including the cyclin-dependent kinase inhibitor p21(waf1/cip1) and the homeobox gene transcription factor HOXA10 are induced at the onset of differentiation. Ectopic expression of either gene results in U937 differentiation. In this paper, we describe a mechanism by which p21 and HOXA10 may act in concert, where HOXA10 can bind directly to the p21 promoter and, together with its trimeric partners PBX1 and MEIS1, activate p21 transcription, resulting in cell cycle arrest and differentiation. These experiments for the first time identify p21 as a selective target for a HOX protein and link the differentiative properties of a transcription factor and a cell cycle inhibitor.
Assuntos
Diferenciação Celular/genética , Ciclinas/genética , Proteínas de Ligação a DNA/metabolismo , Células Mieloides/citologia , Linhagem Celular , Inibidor de Quinase Dependente de Ciclina p21 , Ciclinas/metabolismo , Proteínas de Ligação a DNA/genética , Fase G1/genética , Proteínas Homeobox A10 , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Proteína Meis1 , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Fator de Transcrição 1 de Leucemia de Células Pré-B , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Transcrição Gênica , Ativação TranscricionalRESUMO
Nuclear receptors modulate the transcription of genes in direct response to small lipophilic ligands. Binding to ligands induces conformational changes in the nuclear receptors that enable the receptors to interact with several types of cofactor that are critical for transcription activation (transactivation). We previously described a distinct set of ligand-dependent proteins called DRIPs, which interact with the vitamin D receptor (VDR); together, these proteins constitute a new cofactor complex. DRIPs bind to several nuclear receptors and mediate ligand-dependent enhancement of transcription by VDR and the thyroid-hormone receptor in cell-free transcription assays. Here we report the identities of thirteen DRIPs that constitute this complex, and show that the complex has a central function in hormone-dependent transactivation by VDR on chromatin templates. The DRIPs are almost indistinguishable from components of another new cofactor complex called ARC, which is recruited by other types of transcription activators to mediate transactivation on chromatin-assembled templates. Several DRIP/ARC subunits are also components of other potentially related cofactors, such as CRSP, NAT, SMCC and the mouse Mediator, indicating that unique classes of activators may share common sets or subsets of cofactors. The role of nuclear-receptor ligands may, in part, be to recruit such a cofactor complex to the receptor and, in doing so, to enhance transcription of target genes.
Assuntos
Proteínas Nucleares/fisiologia , Receptores de Calcitriol/fisiologia , Transativadores , Fatores de Transcrição , Ativação Transcricional , Sequência de Aminoácidos , Animais , Proteínas de Transporte/fisiologia , Cromatina/fisiologia , Clonagem Molecular , Drosophila , Células HeLa , Humanos , Ligantes , Substâncias Macromoleculares , Complexo Mediador , Subunidade 1 do Complexo Mediador , Camundongos , Dados de Sequência Molecular , Proteínas Nucleares/química , Homologia de Sequência de AminoácidosRESUMO
The hormonal form of vitamin D, 1,25-dihydroxyvitamin D3 [1, 25(OH)2D3], is a potent inhibitor of cellular proliferation as well as an inducer of differentiation of myeloid leukemic cells to macrophages. We have previously reported that a number of genes are upregulated by 1,25(OH)2D3 during myeloid differentiation, including the cyclin-dependent kinase (CDK) inhibitors p21, p27, 15, and p18, suggesting that cell cycle arrest and differentiation are tightly linked processes. We further explore here the relationship between growth inhibition and differentiation. We report that, upon 1, 25(OH)2D3 treatment, U937 cells exhibited an early proliferative burst followed by growth inhibition and subsequent differentiation. Although CDK levels remain constant throughout, this transient increase in proliferation was accompanied by increases in cyclin A, D1, and E protein levels. p21 and p27 levels were also elevated during both the proliferative burst and subsequent inhibition of cell growth. Ectopic overexpression of p21 and/or p27 in U937 cells, in the absence of hormone, resulted in an induction of the expression of monocyte/macrophage-specific markers, whereas overexpression of p15 and p18 had no effect, suggesting that a subset of CDK inhibitors are important for both growth arrest and differentiation and that an early increase in proliferation is somehow a prerequisite for subsequent differentiation. However, no such biphasic behavior was detected in cells that are growth inhibited by 1,25(OH)2D3 but do not differentiate, such as MCF-7 cells. Taken together, these results indicate that both growth stimulation and subsequent inhibition precede differentiation and involve induction of both cyclins and p21 and p27, whereas cell cycle arrest of differentiated cells can be achieved simply by elevations in CDK inhibitors.