FIG. 6.
Binding of SOX proteins to the Col2a1 enhancer. (A) WT and mutant sequences of the COL2C2 enhancer. To designate COL2C2 mutations, we followed the nucleotide numbering of Lefebvre et al. (32). The C2M(9-14) mutation is the same as C2M in reference 2, which abolishes SOX binding. In C2M(SC), the SOX site was converted to the SOX consensus sequence. In the mutant sequences, altered bases are shaded. Lowercase letters indicate the base sequences added to generate BglII and BamHI restriction sites. (B) Comparison of binding of SOX1, SOX2, and SOX9 to COL2C2 sequences. Gel mobility shift assays were performed with either unprogrammed TNT lysate (−) or S-Tag–SOX proteins (Fig. 2), using the indicated probes. SOX9 bound to the COL2C2 WT and M(1-4) sequences more efficiently than did SOX1 or SOX2. Alteration to C2M(SC) resulted in binding of SOX1 and SOX2 comparable to that of SOX9.