UNLABELLED

Setleis syndrome (SS) is a rare focal facial dermal dysplasia caused by recessive mutations in the basic helix-loop-helix (bHLH) transcription factor, TWIST2. Expression microarray analysis showed that the chordin-like 1 () gene is up-regulated in dermal fibroblasts from three SS patients with the Q119X TWIST2 mutation.

METHODS

Putative TWIST binding sites were found in the upstream region of the gene and examined by electrophoretic mobility shift (EMSA) and reporter gene assays.

RESULTS

EMSAs showed specific binding of TWIST1 and TWIST2 homodimers, as well as heterodimers with E12, to the more distal E-boxes. An adjoining E-box was bound by ADD1/SREBP1c. EMSA analysis suggested that TWIST2 and ADD1/SREBP1c could compete for binding. Luciferase () reporter assays revealed that the gene upstream region drives its expression and ADD1/SREBP1c increased it 2.6 times over basal levels. TWIST2, but not the TWIST2-Q119X mutant, blocked activation by ADD1/SREBP1c, but overexpression of TWIST2-Q119X increased gene expression. In addition, EMSA competition assays showed that TWIST2, but not TWIST1, competes with ADD1/SREBP1c for DNA binding to the same site.

CONCLUSIONS

Formation of an inactive complex between the TWIST2 Q119X and Q65X mutant proteins and ADD1/SREBP1c may prevent repressor binding and allow the binding of other regulators to activate gene expression.