Col1a1 Promoter-targeted Expression of p20 CCAAT Enhancer-binding Protein β (C/EBPβ), a Truncated C/EBPβ Isoform, Causes Osteopenia in Transgenic Mice

Abstract

CCAAT enhancer-binding protein (C/EBP) transcription factors regulate adipocyte differentiation, and recent evidence suggests that osteoblasts and adipocytes share a common pluripotent progenitor in bone marrow. However, little is known about the role of C/EBP transcription factors in the control of osteoblast differentiation or function. In this study, the function of C/EBP transcription factors was disrupted in osteoblast lineage cells by overexpressing a naturally occurring dominant negative C/EBP isoform. Expression of FLp20C/EBP␤ was driven by a 3.6-kb Col1a1 promoter/ first intron construct, and four transgenic (TG) mouse lines were established. Northern blotting and reverse transcription-PCR indicated that the transgene was targeted to bone, with lower levels of expression in lung, skin, and adipose tissue. TG mice from two lines showed reduced body weight compared with wild type littermates. All TG lines showed evidence of osteopenia, ranging from mild to severe, as evidenced by reduced trabecular bone volume. Severely affected lines also showed reduced cortical bone width. Dynamic histomorphometry demonstrated an associated decrease in mineral apposition and bone formation rates. Long bones and calvariae of TG mice showed reduced COL1A1 and osteocalcin mRNA levels and increased bone sialoprotein mRNA, consistent with an inhibition of terminal osteoblast differentiation. Ex vivo analysis of primary osteoblast differentiation showed similar effects on marker expression and reduced expression of the mature osteoblast marker Col2.3-green fluorescent protein, demonstrating a cell-autonomous effect of the transgene. These data suggested that C/EBP transcription factors may be important determinants of osteoblast function and bone mass.CCAAT enhancer-binding proteins (C/EBPs) 1 comprise a family of homologous transcription factors characterized by a carboxyl-terminal leucine zipper dimerization domain and an adjacent highly conserved basic DNA binding domain (1, 2). C/EBPs are transcriptional regulators of gene expression, particularly those involved in energy metabolism (3) and immune or inflammatory responses (4 -6). Over the past several years, C/EBPs have been shown to control cellular differentiation in several lineages, including hepatocytes (7), granulocytes (8), macrophages (9), and adipocytes (10). C/EBP regulation of adipogenesis represents the best characterized system to date. Collectively, the data from a number of pluripotent and pre-adipocytic cell lines support a model in which C/EBP␤ and C/EBP␦ activation represents the seminal event in the process, regulating commitment of progenitor cells to the adipocyte lineage (11). Subsequently, C/EBP␤ and C/EBP␦ induce expression of C/EBP␣ and peroxisome proliferator-activated receptor-␥, which in turn direct expression of terminal adipocyte marker genes (12)(13)(14). Work on C/EBP null mice has confirmed that C/EBP␤ and C/EBP␦ regulate commitment of progenitor cells to the adipocyte lineage, whereas loss of C/EBP␣ function interferes with terminal adip...

CCAAT/Enhancer-binding protein homologous protein (CHOP) decreases bone formation and causes osteopenia

“…This is reaffirmed by recent work demonstrating that mice overexpressing a dominant negative isoform of C/EBP β, p20C/EBP β, LIP, under the control of the type I collagen promoter exhibit osteopenia and a phenotype similar to the one we report in CHOP overexpressing mice [45]. These in vivo findings are in agreement with observations demonstrating that, by interacting with Runx-2, C/EBP β can induce osteocalcin transcription and can promote osteoblast differentiation [6,7].…”

Osteopenia in transgenic mice with osteoblast-targeted expression of the inducible cAMP early repressor

“…ICER transgenic mice have phenotypic similarities to mice with an osteoblast-targeted C/EBPβ p20 transgene including a marked decrease in femoral trabecular bone volume. Since p20 can act as a dominant negative molecule, ICER and p20 transgenes may converge on a similar pathway [53]. In our model, ICER is expressed constitutively at supraphysiological levels.…”

Differences in regulation of type I collagen synthesis in primary and passaged hepatic stellate cell cultures: the role of α5β1-integrin

“…Recent data using overexpression of C/EBP␤ dominant negative isoform in bone of the transgenic animals showed a decrease in bone formation and reduced COL1A1 expression, suggesting that the effect of C/EBP␤ on type I collagen transcription may be similar in all type I collagen-producing cells (12).…”