© 1995 American Heart Association, Inc.
Articles |
Involvement of Multiple cis Elements in Basal- and 
-Adrenergic Agonist–Inducible Atrial Natriuretic Factor Transcription
Roles for Serum Response Elements and an SP-1–Like Element
From the Department of Biology and Molecular Biology Institute, San Diego (Calif) State University.
Correspondence to Dr Christopher C. Glembotski, Department of Biology, San Diego State University, San Diego, CA 92182.E-mail cglembotski@sunstroke.sdsu.edu.
Abstract In the present study, cis elements in
the 5'-flanking sequence (FS) of the rat atrial natriuretic
factor (ANF) gene involved in regulating basal and
1-adrenergic–inducible transcription were
investigated. Truncation analyses using ANF-luciferase reporter
constructs transfected into primary neonatal rat cardiac myocytes
showed that an A/T-rich serum response element (SRE) at -114 bp
of the ANF 5'-FS, which bound serum response factor (SRF), was required
for basal and inducible transcription. In constructs composed of 134 bp
of rat ANF 5'-FS driving luciferase (ANF-134Luc), mutations in the SRE
at -114 bp disrupted SRF binding and ANF promoter activity.
However, the same mutations in ANF-638Luc had little effect, suggesting
a collaborating role for more distal sequences, such as the other SRE
in ANF-638 at -406 bp. In ANF-638Luc, mutations in the SRE at
-406 bp that disrupted SRF binding to that site decreased ANF
reporter activity by only 25%; however, mutating both of the SREs
completely blocked 1-adrenergic–inducible activity.
Mutation analyses showed that an 
(SP-1)–like site at
-69 bp, shown previously to confer inducibility in reporters with
134 bp of ANF 5'-FS, was not required in ANF-638Luc. However, double
mutants in the SP-1–like region and either SRE completely blocked
1-adrenergic–inducible ANF promoter activity. These
findings emphasize that no single element is responsible for
1-adrenergic agonist–regulated ANF transcription
but that the SREs at -114 and -406 bp and the SP-1–like
sequence at -69 bp mediate the effect in collaboration.
Key Words: atrial natriuretic factor • 1-adrenergic agonist inducibility • transcription • serum response element • serum response factor
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