Glyceraldehyde-3-phosphate Dehydrogenase Selectively Binds AU-rich RNA in the NAD-binding Region (Rossmann Fold)

doi:10.1074/jbc.270.6.2755

Glyceraldehyde-3-phosphate Dehydrogenase Selectively Binds AU-rich RNA in the NAD-binding Region (Rossmann Fold) (*)

From the (1)Departments of Medicine and
(2)Microbiology, Dartmouth Medical School, Lebanon, New Hampshire 03756 and
the (3)Veterans Administration Medical Center, White River Junction, Vermont 05009

↵^§ Present address: Dept. of Clinical Chemistry, University Medical School, Pécs H-7643, Hungary.

^¶ To whom correspondence should be addressed:
Section of Connective Tissue Diseases, Dept. of Medicine, Dartmouth-Hitchcock Medical Center, Lebanon, NH 03756.
Tel: 603-650-7700; Fax: 603-650-6223.

Abstract

A 36-kDa protein that binds AU-rich RNA was purified from human spleen and identified as glyceraldehyde-3-phosphate dehydrogenase (GAPDH). GAPDH has been previously demonstrated to bind tRNA with high affinity. Competition studies suggested that cytoplasmic GAPDH binds the AU-rich elements (AREs) of lymphokine mRNA 3′-untranslated regions with higher affinity than tRNA. The AUUUA-specific RNA binding activity of GAPDH was inhibited by NAD, NADH, and ATP in a concentration-dependent manner, suggesting that RNA binding of GAPDH might involve the NAD-binding region, or dinucleotide-binding (Rossmann) fold. This hypothesis was supported by experiments that localized RNA binding to the predicted N-terminal 6.8-kDa peptide, known to be involved in the formation of the NAD-binding domain. The direct demonstration of AREspecific binding protein activity localized to the NAD-binding region of GAPDH supports the general concept that enzymes containing this domain may exhibit specific RNA binding activity and play additional roles in nucleic acid metabolism. Finally, cytoplasmic GAPDH was found in the polysomal fraction of T lymphocytes. Thus, the RNA binding specificity of GAPDH as well as its localization within the cell merit its strong consideration as a protein important in the regulation of ARE-dependent mRNA turnover and translation in addition to its well described role in glycolysis.

Footnotes

↵* This work was supported by National Institutes of Health Grants RO1 AI2434 and KO4 AI00910 and by research funds from the Department of Veterans Affairs. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

AREs

AU-rich elements

AUBPs

ARE-specific binding proteins

hnRNP

heterogeneous nuclear ribonucleoprotein

IL-2

interleukin-2

GAPDH

glyceraldehyde-3-phosphate dehydrogenase

CAPS

3-(cyclohexylamino)propanesulfonic acid

Tricine

N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine

PAGE

polyacrylamide gel electrophoresis

UTR

untranslated region

IFN-

interferon-

GM-CSF

granulocyte-macrophage colony-stimulating factor

ptRNA

precursor tRNA.
↵² B. J. Hamilton and W. F. C. Rigby, unpublished observation.
- Received August 3, 1994.
- Revision received October 10, 1994.