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Identification of an osteocalcin gene promoter sequence that binds AP1
Author(s) -
Goldberg Daniella,
Polly Patsie,
Eisman John A.,
Morrison Nigel A.
Publication year - 1996
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/(sici)1097-4644(19960315)60:4<447::aid-jcb2>3.0.co;2-s
Subject(s) - ap 1 transcription factor , osteocalcin , osteoblast , promoter , biology , microbiology and biotechnology , consensus sequence , activator (genetics) , gene , electrophoretic mobility shift assay , transcription factor , transcriptional regulation , gene expression , genetics , biochemistry , peptide sequence , in vitro , alkaline phosphatase , enzyme
Osteoblasts are differentiated cells that produce bone matrix components including the bone‐specific protein osteocalcin. The osteocalcin gene promoter has become a model for understanding how genes are regulated, specifically in osteoblasts. One model for cell‐specific regulation suggests that osteoblast‐expressed genes are regulated through common promoter sequences which bind osteoblast‐specific transcriptional activators. The phenotype suppression model suggests osteoblast‐specific promoters are switched off through the action of the common transcriptional activator AP1. We previously demonstrated that a short sequence element (OSCARE‐2) in the osteocalcin promoter was homologous to a repressive element in the collagen type 1 (α1) promoters. In this paper we use electrophoretic mobility shift (EMS) assays to examine DNA‐protein interactions in the OSCARE‐2 sequence. In EMS assays, OSCARE‐2 binds a complex of proteins, including AP1. This supports the role of AP1 sites in contributing to the regulation of the osteocalcin promoter. Exogenous c‐JUN protein bound to OSCARE‐2 and increasing c‐JUN incubated with nuclear extract amounts caused a progressive increase in a higher‐molecular‐weight complex, consistent with c‐JUN involvement in protein‐protein as well as DNA‐protein interactions. Anti‐c‐FOS antibody was capable of supershifting OSCARE‐2 DNA‐protein complexes produced using osteoblast‐like cell nuclear extracts. In addition, EMS assays of nuclear proteins from osteoblast‐like cells indicated that 1,25 (OH) 2 D 3 ‐inducible proteins are bound to OSCARE‐2. Osteocalcin promoter constructs showed that OSCARE‐2 contributed to the 1,25 (OH) 2 D 3 response, albeit in a minor way. These data support the role of AP1 protein as a regulator of osteoblast‐specific gene expression during osteoblast development. © 1996 Wiley‐Liss, Inc.