CREB-AP1 Protein Complexes Regulate Transcription of the Collagen XXIV Gene (Col24a1) in Osteoblasts

Shizuko Tanaka,Hidekatsu Yoshioka,Francesco Ramirez,Marion K. Gordon,Manuel Koch,Noritaka Matsuo

doi:10.1074/jbc.m509923200

Abstract

Collagen XXIV is a newly discovered and poorly characterized member of the fibril-forming family of collagen molecules, which displays unique structural features of invertebrate fibrillar collagens and is expressed predominantly in bone tissue. Here we report the characterization of the proximal promoter of the mouse gene (Col24a1) and its regulation in osteoblastic cells. Using well characterized murine models of osteoblast differentiation, we found that the Col24a1 gene is activated sometime before onset of the late differentiation marker osteocalcin. Additional analyses revealed that Col24a1 produces equal amounts of two alternatively spliced products with different 5'-untranslated sequences that originate from distinct transcriptional start sites. Cell transfection experiments in combination with DNA binding assays demonstrated that Col24a1 promoter activity in ROS17/2.8 osteosarcoma cells is under the control of an upstream cis-acting element, which is shared by both transcripts and is recognized by specific combinations of c-Jun, CREB1, ATF1, and ATF2 dimers. Consistent with these results, overexpression of c-Jun, ATF1, ATF2, or CREB1 in transiently transfected osteoblastic cells stimulated transcription from reporter gene constructs driven by the Col24a1 promoter to different degrees. Moreover, chromatin immunoprecipitation experiments showed that these nuclear factors bind the same upstream sequence of the endogenous Col24a1 gene. Collectively these data provide new information about transcriptional control of collagen fibrillogenesis, in addition to implicating for the first time CREB-AP1 protein complexes in the regulation of collagen gene expression in osteoblasts.

Highlights

Vertebrate collagens represent a very large superfamily of extracellular proteins that impart specific physical properties to the connective tissue of virtually every organ system [1,2,3]
This study extends our knowledge of the transcriptional regulation of collagen fibrillogenesis, in addition to implicating for the first time CREB-AP1 protein complexes in the expression of a fibrillar collagen gene in osteoblasts
The present study demonstrated that Col24a1 is a marker of late osteoblast differentiation which is positively regulated by the binding of specific combinations of CREB-AP1 proteins to an upstream cis-acting element, which is shared by the two alternative promoters of the gene

Summary

Introduction

Vertebrate collagens represent a very large superfamily of extracellular proteins that impart specific physical properties to the connective tissue of virtually every organ system [1,2,3]. Several studies have identified DNA cis-acting elements and nuclear trans-acting factors that regulate cartilage-specific expression of the collagen II and XI genes (29 –32).

Results

Conclusion