RESUMEN
More than 40 years after the discovery of vasoactive intestinal peptide (VIP), its transcriptome in the immune system has still not been completely elucidated. In an attempt to understand the biological role of this neuropeptide in immunity, we chose CD4 T cells as a cellular system. Agilent Mouse Whole Genome microarrays were hybridized with fluorescently labeled total RNA isolated from resting CD4 T cells cultured +/-10(-7)M VIP for 5h or PMA/ionomycin activated CD4 T cells cultured +/-10(-7)M VIP for 5h. These VIP-regulated transcriptomes were analyzed by Significance Analysis of Microarrays (SAM) and Ingenuity Pathway Analysis (IPA) software to identify relevant signaling pathways modulated by VIP in the absence and presence of T cell activation. In resting CD4 T cells, VIP-modulated 368 genes, ranging from 3.49 to -4.78-fold. In the PMA/ionomycin activated CD4 T cells, 326 gene expression levels were changed by VIP, ranging from 2.94 to -1.66-fold. IPA analysis revealed that VIP exposure alters cellular function through EGFR signaling in resting CD4 T cells, and modulates immediate early genes, Fos and CREM/ICER, in activated CD4 T cells. These gene expression changes are suggested to explain at a molecular level how VIP can regulate T cell homing to the gut and induce regulatory T cell generation.
Asunto(s)
Linfocitos T CD4-Positivos/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/genética , Transducción de Señal/efectos de los fármacos , Péptido Intestinal Vasoactivo/farmacología , Animales , Linfocitos T CD4-Positivos/efectos de los fármacos , Femenino , Redes Reguladoras de Genes , Masculino , Ratones , Ratones Endogámicos C57BL , Análisis de Secuencia por Matrices de Oligonucleótidos , Fenotipo , Receptores de Péptido Intestinal Vasoactivo/genética , Receptores de Péptido Intestinal Vasoactivo/metabolismo , Reproducibilidad de los Resultados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Acetato de Tetradecanoilforbol/farmacologíaRESUMEN
Hematopoietic defects in HOXA9(-/-) mice demonstrate a key role for this homeoprotein in blood cell development. Conversely, enforced HOXA9 expression is leukemogenic in mice, and HOXA9 is frequently activated in human acute myeloid leukemia (AML). Although HOXA9 is thought to function as a transcription factor, few downstream targets have been identified. We searched for early HOXA9 target genes by using a transient overexpression strategy in 3 hematopoietic cell lines (2 myeloid, 1 lymphoid). cDNA microarray analyses identified 220 genes whose expression was modulated at least 2-fold. Expression signatures in myeloid and lymphoid cells demonstrated that HOXA9 functions as both an activator and repressor of a variety of genes in cell-specific patterns suggesting that the transcriptional effects of HOXA9 are largely dependent on the cell context. Transient transcription assays and target gene expression patterns in HOXA9(-/-) marrow cells imply that we have identified direct physiologic targets. Many target genes are expressed in CD34+ stem cells or are members of gene families involved in proliferation or myeloid differentiation. Expression of 14 HOXA9 target genes correlated with high-level HOXA9 expression in primary AML. These data suggest that many genes identified in this survey may mediate the biologic effects of HOXA9 in normal and leukemic hematopoiesis.