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Originally published In Press as doi:10.1074/jbc.M009352200 on October 18, 2000

J. Biol. Chem., Vol. 276, Issue 2, 1026-1033, January 12, 2001
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GATA-4 and Serum Response Factor Regulate Transcription of the Muscle-specific Carnitine Palmitoyltransferase I beta  in Rat Heart*

Meredith L. MooreDagger , Guo-Li WangDagger , Narasimhaswamy S. Belaguli§, Robert J. Schwartz§, and Jeanie B. McMillinDagger

From the Dagger  Department of Pathology and Laboratory Medicine, Medical School, University of Texas-Houston Health Science Center and the § Department of Cell Biology, Baylor College of Medicine, Houston, Texas 77030

Transcriptional regulation of nuclear encoded mitochondrial proteins is dependent on nuclear transcription factors that act on genes encoding key components of mitochondrial transcription, replication, and heme biosynthetic machinery. Cellular factors that target expression of proteins to the heart have been well characterized with respect to excitation-contraction coupling. No information currently exists that examines whether parallel transcriptional mechanisms regulate nuclear encoded expression of heart-specific mitochondrial isoforms. The muscle CPT-Ibeta isoform in heart is a TATA-less gene that uses Sp-1 proteins to support basal expression. The rat cardiac fatty acid response element (-301/-289), previously characterized in the human gene, is responsive to oleic acid following serum deprivation. Deletion and mutational analysis of the 5'-flanking sequence of the carnitine palmitoyltransferase Ibeta (CPT-Ibeta ) gene defines regulatory regions in the -391/+80 promoter luciferase construct. When deleted or mutated constructs were individually transfected into cardiac myocytes, CPT-I/luciferase reporter gene expression was significantly depressed at sites involving a putative MEF2 sequence downstream from the fatty acid response element and a cluster of heart-specific regulatory regions flanked by two Sp1 elements. Each site demonstrated binding to cardiac nuclear proteins and competition specificity (or supershifts) with oligonucleotides and antibodies. Individual expression vectors for Nkx2.5, serum response factor (SRF), and GATA4 enhanced CPT-I reporter gene expression 4-36-fold in CV-1 cells. Although cotransfection of Nkx and SRF produced additive luciferase expression, the combination of SRF and GATA-4 cotransfection resulted in synergistic activation of CPT-Ibeta . The results demonstrate that SRF and the tissue-restricted isoform, GATA-4, drive robust gene transcription of a mitochondrial protein highly expressed in heart.

* This work was supported by National Institutes of Health Grants RO1 HL38863 (to J. B. M.) and PO1-HL49953 (to R. J. S.), and by United States Department of Agriculture Grant ARS6250-6100 (to R. J. S.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

To whom correspondence should be addressed: Dept. of Pathology and Laboratory Medicine, Medical School, University of Texas-Houston Health Science Center, 6431 Fannin, Houston, TX 77030. Tel.: 713-500-5335; Fax: 713-500-0730; E-mail: jeanie.b.mcmillin@uth.tmc.edu.

Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.
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