Doubly Truncated FosB Isoform (Δ2ΔFosB) Induces Osteosclerosis in Transgenic Mice and Modulates Expression and Phosphorylation of Smads in Osteoblasts Independent of Intrinsic AP-1 Activity

Abstract

ABSTRACT:Introduction: Activator protein (AP)-1 family members play important roles in the development and maintenance of the adult skeleton. Transgenic mice that overexpress the naturally occurring ⌬FosB splice variant of FosB develop severe osteosclerosis. Translation of ⌬fosb mRNA produces both ⌬FosB and a further truncated isoform (⌬2⌬FosB) that lacks known transactivation domains but, like ⌬FosB, induces increased expression of osteoblast marker genes. Materials and Methods:To test ⌬2⌬FosB's ability to induce bone formation in vivo, we generated transgenic mice that overexpress only ⌬2⌬FosB using the enolase 2 (ENO2) promoter-driven bitransgenic Tet-Off system. Results: Despite ⌬2⌬FosB's failure to induce transcription of an AP-1 reporter gene, the transgenic mice exhibited both the bone and the fat phenotypes seen in the ENO2-⌬FosB mice. Both ⌬FosB and ⌬2⌬FosB activated the BMP-responsive Xvent-luc reporter gene and increased Smad1 expression. ⌬2⌬FosB enhanced BMP-induced Smad1 phosphorylation and the translocation of phospho-Smad1 (pSmad1) to the nucleus more efficiently than ⌬FosB and showed a reduced induction of inhibitory Smad6 expression. Conclusions: ⌬FosB's AP-1 transactivating function is not needed to induce increased bone formation, and ⌬2⌬FosB may act, at least in part, by increasing Smad1 expression, phosphorylation, and translocation to the nucleus.

Antagonistic Regulation of Cell-Matrix Adhesion by FosB and ΔFosB/Δ2ΔFosB Encoded by Alternatively Spliced Forms of fosB Transcripts

“…We thus conclude that ⌬FosB and/or ⌬2⌬FosB positively regulate cell-matrix adhesion as well as TGF-␤1 signaling, whereas FosB negatively regulates them. Recently, it has been reported that ⌬2⌬FosB and to a lesser extent ⌬FosB modulate the expression and phosphorylation of Smads independent of intrinsic AP-1 activity (Sabatakos et al, 2008), confirming that both ⌬FosB and ⌬2⌬FosB play important roles in the up-regulation of TGF-␤1 signaling, as we observed in the present study.…”

“…It has been shown that expression of the Tgfb1i1 gene is regulated by RAR as well as by TGF-␤1, both of which can modulate AP-1 function (Liberati et al, 1999;Zhou et al, 1999;Suzukawa and Colburn, 2002;Zhuang et al, 2003), suggesting that expression of Tgfb1i1 is indirectly regulated by FosB or ⌬FosB/⌬2⌬FosB through their interaction with steroid receptors or Smad proteins, as reported recently (Sabatakos et al, 2008). Furthermore, it has been shown that Hic5/ARA55 itself negatively regulates Smad3 signaling (Wang et al, 2005).…”

“…In contrast, genes whose expression was different between the two are considered to be regulated by ⌬FosB/⌬2⌬FosB. These results are unexpected in that they imply that expression of a substantial number of genes is directly or indirectly regulated by ⌬FosB and that expression of a much smaller number of genes is regulated by FosB, because FosB is a more potent transactivator than ⌬FosB or ⌬2⌬FosB (Nakabeppu and Nathans, 1991;Sabatakos et al, 2008).…”

Direct Transcriptional Repression of Zfp423 by Zfp521 Mediates a Bone Morphogenic Protein-Dependent Osteoblast versus Adipocyte Lineage Commitment Switch

“…To examine BMP2 responses, cells were maintained in 0.5% fetal bovine serum and treated with 100 ng/ml BMP2 or vehicle 16 h prior to assay. Flag-Ebf1, Smad1, and Smad4 constructs were described previously (22,32). Zfp521 deletion constructs (21) were kind gifts from D. Kamiya and Y. Sasai.…”

Zfp521 controls bone mass by HDAC3-dependent attenuation of Runx2 activity

“…First we generated mice with a heterozygous germline deletion of Zfp521 (Zfp521 We sought to further investigate the Zfp521-mediated antagonism of Runx2 activity by generating bitransgenic mice in which overexpression of Zfp521 was regulated by the enolase 2 (ENO2) promoter (ENO2-tTA;TetOp-Zfp521 mice, hereafter referred to as Zfp521 Tg mice). We successfully used this bitransgenic system in earlier osteoblast-related work (Sabatakos et al, 2000(Sabatakos et al, , 2008Rowe et al, 2009) and selected it for this study because the ENO2 promoter is active in both early osteoblasts (Fig. S1 A) and in more mature osteoblasts in the long bones of adult mice (Fig.…”