Title

The role of CREM/ICER transcription factors in bone

Date of Completion

January 2006

Keywords

Biology, Molecular

Degree

Ph.D.

Abstract

Crem, the cyclic AMP response element modulator, is an important component of cAMP-mediated signal transduction and couples extracellular signals to gene regulation. The Crem gene has 4 different promoters (P1 through P4), which drive the expression of different Crem isoforms. An intronic promoter (P2) near the 3' end of the Crem gene directs the transcription of the inducible cyclic AMP early repressor, ICER, which is cAMP-inducible and consists primarily of the basic leucine zipper motif of CREM. The objective of this thesis was to study the role of CREM/ICER in bone. ^ We found that osteoblasts expressed at least 15 Crem transcripts initiated from the P1 promoter, including 7 novel transcripts. CREM-x contains a new exon termed L. Therefore, Crem gene expression in osteoblasts is unique and complex. Crem deficiency in mice altered the response of bone to intermittent PTH treatment such that osteoclastogenesis was increased. Thus, the Crem gene may specify the anabolic response to intermittent PTH treatment by restraining PTH-induced osteoclastogenesis. ^ Crem deficiency in osteoblasts did not alter the time course of c-Fos, IL-6 and COX-2 expression in response to 10 μm forskolin or 10 nM PTH, suggesting that CREM/ICER may not play a significant role in the attenuation of cAMP inducible gene expression in osteoblasts. Overexpression of ICER in osteoblasts of transgenic mice decreased the induction of c-Fos and IL-6 in response to forskolin (FSK) but increased the induction of Cox-2 suggesting that ICER may regulate cAMP inducible gene expression differentially. Transgenic ICER decreased the activity of a Cox-2 luciferase reporter in osteoblasts. ^ A future direction of this work includes the generation of osteoblast specific CREM/ICER gene ablation using the Cre/loxP recombination system. As a first step, Cre recombinase has been targeted to cells of the osteoblast lineage with 2.3 kb (Col2.3-Cre) and 3.6 kb (Col3.6-Cre) fragments of the rat Col1a1 promoter and characterized by breeding with ROSA26 (R26R) mice. The spatial pattern of 13-gal staining was more restricted in bone and in bone marrow stromal cultures established from Col2.3-Cre;R26R mice. Col2.3-Cre and Col3.6-Cre transgenic mice may be useful for conditional gene targeting.^