GENE REGULATION OF HUMAN INTERLEUKIN-9 (IL-9)

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The cDNA encoding human IL-9 was isolated from a HTLV-I transformed human T cell line through mammalian cell expression cloning method based on its ability to stimulate the proliferation of a factor-dependent human megakaryocytic leukemic line. The newly isolated cytokine has subsequently been shown to support the growth of human erythroid progenitors in vitro. The long term objective of this proposal is to understand the role of human IL-9 in regulating hematopoiesis at the molecular and cellular level. The proposed work specifically aims to (1) determine the regulatory mechanism of human IL-9 gene expression at the molecular and cellular level. This will be achieved by (i) analyzing the cis-elements required for IL-9 gene expression through measuring the promoter activities of plasmid constructs containing various portions of IL-9 5'-flanking region upstream from a luciferase reporter gene in various cell types under different stimuli, (ii) examining the transacting factors responsible for IL-9 gene expression through DNase I footprinting, gel retardation , methylation interference assay and site-specific mutagenesis and (iii) initiating the characterization and purification of the transacting factors and molecular cloning for the genes encoding these proteins; (2) evaluate the possible mechanism of human IL-9 expression in HTLV-I associated leukemia. We will test whether the constitutive expression of human IL-9 in HTLV-transformed cells is due to (i) the transactivation mechanism elicited by HTLV tax protein or tax-induced proteins or (ii) the insertion/activation mechanism through viral integration. The study of gene regulation of this new cytokine should lead to a better understanding of the regulatory mechanism of human IL-9 production in the complicated cytokine network and the possible involvement of human IL-9 in leukemogenesis.