Alpha-Globin expression: Post transcriptional mechanisms

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High-level stability of globin mRNAs is a major determinant of hemoglobin synthesis and erythrocyte function. The basis for selective stabilization of globin mRNAs during erythroid differentiation remains poorly understood. Our laboratory is using human ot-globin mRNA as a model for the study of this problem. Genetic, biochemical, and in vivo expression studies carried out over the present funding period point to a central role for a sequence-specific 3'UTR RNA-protein (RNP) complex ('c_ complex') in stabilizing ct- globin mRNA. Inactivation of the c_-complex by mutation of the C-rich binding motif or by blocking the binding of the ctCP protein results in an incremental loss of ot-globin mRNA stability. This loss of stability can be fully restored by artificially tethering ctCP to the 3'UTR. c_CPs are broadly distributed in tissues, suggesting that an erythroid- restricted role of the a-complex is dictated by specific modifications to c_CP or to interacting RNP components. The major ctCP isoforms are differentially localized in the nucleus and cytoplasm. Evidence suggests that the cytoplasmic role of etCPs in ct-globin mRNA stabilization is complemented by separate nuclear function(s) involved in enhancement of c_-globin mRNA processing. The pathways involved in selective stabilization of human c_-globin mRNA and the interrelationships between nuclear and cytoplasmic functions of aCPs in c_-globin gene expression will be explored in the proposed studies. Aim I. Identify interactions at the a complex that mediate ct-globin mRNA stabilization. Aim II. Define the mechanism(s) of _-globin mRNA stabilization and how a-globin mRNA evades decay in erythroid cells. Aim III. Determine how ctCPs enhance nuclear processing of ct-globin transcripts and how these nuclear events integrate with aCP-mediated cytoplasmic controls. These studies will extend our prior work on ct-globin gene expression, define novel pathways of mRNA decay, and establish a paradigm for coordinated nuclear and cytoplasmic post-transcriptional controls in erythroid gene expression.