Expression and function of Bapx1 during chick limb development

The homeobox-containing transcription factor Bapx1 (also known as Nkx3.2) is crucial for development of the axial skeleton and parts of the chondrocranium. Here we describe the detailed expression of Bapx1 during chick limb development and show that in contrast to its expression in the axial skeleton, Bapx1 is expressed after the commitment to chondrogenesis. Bapx1 is initially expressed throughout the developing skeletal elements prior to the overt differentiation of the distinct chondrogenic layers. Once distinct layers (proliferating, prehypertrophic and hypertrophic) have formed, Bapx1 expression is restricted to the proliferating chondrocytes. Bapx1 transcripts are excluded from the articular cartilage. A second homeobox-containing transcription factor, Barx1, is expressed in a complementary fashion in the developing joint and articular cartilage. Interestingly, in vitro functional analyses showed that Bapx1 overexpression in micromass cultures increased both matrix production and nodule number suggesting that Bapx1 is sufficient to promote chondrogenesis in the limb. In contrast, Barx1 had the opposite effect on nodule number suggesting that it has an inhibitory effect on chondrogenic initiation consistent with its expression in the developing joint. A slight increase in matrix levels was also observed consistent with its expression in the articular chondrocytes. Finally, we show that Bapx1 is also expressed in the soft tissues such as the developing tendons, muscle sheaths and surrounding mesenchyme, and therefore may have additional as yet uncharacterized roles in limb morphogenesis.