Identification of Pax6-binding
sites of the c-maf gene. (A) Schematic
representation of the probes used for EMSA and footprinting analyses.
(B) EMSA of c-maf promoter fragments
with Pax6–MBP fusion protein. The terminally labeled fragments
indicated were bound with Pax6–MBP protein (100 ng) and
analyzed as described in Materials and Methods. (C)
Footprinting analysis of c-maf promoter fragments.
Probe A, E or F was mixed with Pax6–MBP protein (1 µg), treated with DNase I and analyzed
by denatured polyacrylamide gel electrophoresis. The same probes,
modified and digested by the Maxam–Gilbert method, were used
as makers (M). (+) and (–) indicate with or without
Pax6–MBP protein, respectively. Footprinting regions are
indicated by vertical bars and lined under the sequences. (D)
Pax6-binding sequences. Pax6 paired domain binding consensus sequence
obtained by PCR-based oligonucleotide selection (33) and that of
the promoter region of the ζ-crystallin
gene (28) are aligned together with binding sequences of the c-maf promoter.