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.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37HL065449-14
Application #
8502515
Study Section
Special Emphasis Panel (NSS)
Program Officer
Qasba, Pankaj
Project Start
2000-09-05
Project End
2014-06-30
Budget Start
2013-07-01
Budget End
2014-06-30
Support Year
14
Fiscal Year
2013
Total Cost
$404,307
Indirect Cost
$147,960
Name
University of Pennsylvania
Department
Genetics
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Kini, Hemant K; Kong, Jian; Liebhaber, Stephen A (2014) Cytoplasmic poly(A) binding protein C4 serves a critical role in erythroid differentiation. Mol Cell Biol 34:1300-9
Ghanem, Louis R; Chatterji, Priya; Liebhaber, Stephen A (2014) Specific enrichment of the RNA-binding proteins PCBP1 and PCBP2 in chief cells of the murine gastric mucosa. Gene Expr Patterns 14:78-87
Harper, Sandra L; Sriswasdi, Sira; Tang, Hsin-Yao et al. (2013) The common hereditary elliptocytosis-associated *-spectrin L260P mutation perturbs erythrocyte membranes by stabilizing spectrin in the closed dimer conformation. Blood 122:3045-53
Ji, Xinjun; Wan, Ji; Vishnu, Melanie et al. (2013) *CP Poly(C) binding proteins act as global regulators of alternative polyadenylation. Mol Cell Biol 33:2560-73
Natsuizaka, Mitsuteru; Naganuma, Seiji; Kagawa, Shingo et al. (2012) Hypoxia induces IGFBP3 in esophageal squamous cancer cells through HIF-1*-mediated mRNA transcription and continuous protein synthesis. FASEB J 26:2620-30
Ji, Xinjun; Kong, Jian; Liebhaber, Stephen A (2011) An RNA-protein complex links enhanced nuclear 3' processing with cytoplasmic mRNA stabilization. EMBO J 30:2622-33
Vishnu, Melanie R; Sumaroka, Marina; Klein, Peter S et al. (2011) The poly(rC)-binding protein alphaCP2 is a noncanonical factor in X. laevis cytoplasmic polyadenylation. RNA 17:944-56
Kini, Hemant K; Vishnu, Melanie R; Liebhaber, Stephen A (2010) Too much PABP, too little translation. J Clin Invest 120:3090-3
Waggoner, Shelly A; Johannes, Gregg J; Liebhaber, Stephen A (2009) Depletion of the poly(C)-binding proteins alphaCP1 and alphaCP2 from K562 cells leads to p53-independent induction of cyclin-dependent kinase inhibitor (CDKN1A) and G1 arrest. J Biol Chem 284:9039-49
Young, Regina M; Wang, Shang-Jui; Gordan, John D et al. (2008) Hypoxia-mediated selective mRNA translation by an internal ribosome entry site-independent mechanism. J Biol Chem 283:16309-19

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