(D) Transactivation assays were performed in COS1 cells transiently cotransfected with the IL4 promoter cloned into pGL3 vector reporter construct (kindly provided by Michael Lohoff, University of Marburg, Marburg, Germany) alone (black bar) or together with wild-type MAF (white bar) or each of the disease-causing MAF mutants (blue and red bars) (1:1 ratio), and 1:10 of Renilla luciferase control vector DNA (pRL-Act Renilla). After transfection (24 hr), firefly and Renilla luciferase activities were measured by the Dual Luciferase Reporter Assay System (Promega). Normalized luciferase activity (mean ± SD) of six experiments performed is reported as fold increase relative to cells not expressing exogenous MAF. p values were calculated using two-tailed Student’s t test. ∗, ∗∗, and ∗∗∗ indicate p < 0.05, p < 0.01, and p < 0.001, respectively. Protein levels of wild-type and disease-causing mutant MAF proteins were evaluated by immunoblotting with anti-MAF and anti-actin antibodies (lower panels).