Endogenous Msx1 antisense transcript: In vivo and in vitro evidences, structure, and potential involvement in skeleton development in mammals

Msx1 is a key factor for the development of tooth and craniofacial skeleton and has been proposed to play a pivotal role in terminal cell differentiat ion. In this paper, we demonstrated the presence of an endogenous Msx1 anti sense RNA (Msx1-AS RNA) in mice, rats, and humans. In site analysis reveale d that this RNA is expressed only in differentiated dental and bone cells w ith an inverse correlation with Msx1 protein. These in vivo data and overex pression of Msx1 sense and AS RNA in an odontoblastic cell line (MO6-G3) sh owed that the balance between the levels of the two Msx1 RNAs is related to the expression of Msx1 protein. To analyze the impact of this balance in t he Msx-Dlx homeoprotein pathway, we analyzed the effect of Msx1, Msx2, and Dlx5 overexpression on proteins involved in skeletal differentiation. We sh owed that the Msx1-AS RNA is involved in crosstalk between the Msx-Dlx path ways because its expression was abolished by Dlx5. Msx1 was shown to down-r egulate a master gene of skeletal cells differentiation, Cbfa1. All these d ata strongly suggest that the ratio between Msx1 sense and antisense RNAs i s a very important factor in the control of skeletal terminal differentiati on. Finally, the initiation site for Msx1-AS RNA transcription was located by primer extension in both mouse and human in an identical region, includi ng a consensus TATA box, suggesting an evolutionary conservation of the AS RNA-mediated regulation of Msx1 gene expression.