A ubiquitous factor (HF-1a) and a distinct muscle factor (HF-1b/MEF-2) form an E-box-independent pathway for cardiac muscle gene expression.
Recent studies have identified a conserved 28-bp element (HF-1) within the rat cardiac MLC-2 gene which confers cardiac muscle-specific and inducible expression during myocardial cell hypertrophy. Utilizing a combination of independent experimental approaches, this study characterizes two cardiac nuclear factors which bind to HF-1, a ubiquitous factor (HF-1a), and an A + T-rich binding factor (HF-1b) which is preferentially expressed in differentiated cardiac and skeletal muscle cells. The HF-1a binding site is located in a core region of the 28-bp conserved element, immediately upstream from the A + T-rich HF-1b site, which is homologous to the MEF-2 site found in a number of muscle genes. By a number of separate criteria (gel mobility shift, competition, and mutagenesis studies), HF-1b and MEF-2 appear to be indistinguishable and thus are either identical or closely related muscle factors. Transient assays of luciferase reporter genes containing point mutations throughout the 28-bp HF-1 regulatory element document the importance of both the HF-1a and HF-1b sites in transient assays in ventricular muscle cells. In the native 250-bp MLC-2 promoter fragment, mutations in the single E box had little effect on cardiac muscle specificity, while point mutations in either the HF-1a or HF-1b binding site significantly reduced promoter activity, underscoring the importance of both the HF-1a and HF-1b sites in the transcriptional activation of this cardiac muscle gene. Thus, this study provides evidence that a novel, ubiquitous factor (HF-1a) and a muscle factor (HF-1b/MEF-2) can form a novel, E-box-independent pathway for muscle-specific expression in ventricular cardiac muscle cells.
Navankasattusas
S
S
Department of Medicine, University of California, San Diego, La Jolla 92093-0613.
Zhu
H
H
Garcia
A V
AV
Evans
S M
SM
Chien
K R
KR
D000818:Animals; D001483:Base Sequence; D006332:Cardiomegaly / Q000378:metabolism; D002454:Cell Differentiation; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism* / Q000473:pathology; D051381:Rats; D011993:Recombinant Fusion Proteins; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D014018:Tissue Distribution
D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins