Fig. 5. Retroviral expression of Barx2 and HD-BBR-C proteins in limb bud micromass cultures increases chondrogenic differentiation. (A) Three different Barx2 proteins encoded in retroviral vectors: the full-length Barx2 protein (Barx2) that contains the homeodomain (HD), Barx basic region (BBR) and an acidic activation domain (AD); the HD-BBR protein contains only the DNA-binding region of Barx2; and the HD-BBR-C protein, which contains the DNA-binding region as well as the C-terminal activation domain. (B) Expression of Barx2 or HD-BBR-C proteins, but not HD-BBR protein, induced chondrogenesis in limb bud micromass cultures as indicated by Alcian Blue staining. (C) Quantitative analyses of Alcian Blue incorporation in retroviral infection experiments; n, number of experiments. (D) Micromass cultures prepared from proximal limb bud mesenchyme show no response to overexpression of control, Barx2, HD-BBR-C or HD-BBR proteins, whereas distal limb bud cultures show a fivefold increase in Alcian Blue staining upon infection with the Barx2 retroviral construct; n, number of experiments.