FIGURE 7.

Site-directed mutagenesis analysis of region II. A, schematic representation of site-directed mutagenesis of region II. B, the wild type (WTII) hNT/N promoter fragment (−106 to −77) and its mutants (M5–M8) were radiolabeled and incubated with BON cell nuclear extracts (10 μg), and was performed as described under “Experimental Procedures.” C, competitive binding analysis by EMSA with BON nuclear extracts (10 μg) and unlabeled WTII and mutant probes M5–M8 (at 200-fold molar excesses) as competitors. D, the −175 to +26 region of the hNT/N promoter and its mutants (M5–M8) were cloned into the luciferase reporter plasmid, pXP1, and transiently transfected into BON cells. Luciferase activities are expressed as a percentage of the wild type (−175 to +26) fusion plasmid and are the mean ± S.D. of four separate transfections after normalization to β-galactosidase expression.