FUNCTIONAL-ROLE OF A CONFORMATIONALLY FLEXIBLE HOMOPURINE HOMOPYRIMIDINE DOMAIN OF THE ANDROGEN RECEPTOR GENE PROMOTER INTERACTING WITH SP1AND A PYRIMIDINE SINGLE-STRAND DNA-BINDING PROTEIN/
S. Chen et al., FUNCTIONAL-ROLE OF A CONFORMATIONALLY FLEXIBLE HOMOPURINE HOMOPYRIMIDINE DOMAIN OF THE ANDROGEN RECEPTOR GENE PROMOTER INTERACTING WITH SP1AND A PYRIMIDINE SINGLE-STRAND DNA-BINDING PROTEIN/, Molecular endocrinology, 11(1), 1997, pp. 3-15
The androgen receptor (AR) gene promoter does not contain the TATA or
CAAT box, but it contains a long (similar to 90-bp) homopurine/homopyr
imidine (pur/pyr) stretch immediately upstream of the Sp1-binding GC b
ox site. This pur/pyr stretch is conserved at the same proximal positi
on in the rat, mouse, and human AR gene promoters. Mutation of this re
gion results in a 3-fold decline in promoter activity, indicating an i
mportant regulatory function. Examination of the conformational state
of the AR pur/pyr region with the single-strand-specific S1 nuclease s
howed that it is capable of forming a non-B DNA structure involving un
paired single strands. Fine mapping of the S1-sensitive site revealed
an unsymmetric cleavage pattern indicative of an intramolecular triple
helical H-form DNA conformation. Electrophoretic mobility shift analy
ses showed that the pur/pyr region of the AR promoter can bind a novel
pyrimidine single-strand-specific protein (ssPyrBF) and also a double
-strand DNA-binding protein. Both oligonucleotide cross-competition an
d antibody supershift experiments established that the double-strand b
inding protein is equivalent to Sp1. Deoxyribonuclease I (DNase I) foo
tprinting analysis showed multiple Sp1-binding to the pur/pyr site and
a weaker Sp1 interaction to this region compared with the adjacently
located GC box, where Sp1 functions to recruit the TFIID complex. Thes
e results suggest that the pur/pyr domain of the AR gene can serve to
attract additional Sp1 molecules when it exists in the double-stranded
B-DNA conformation. However, binding of ssPyrBF and the resultant sta
bilization of the non-B DNA structure is expected to prevent its inter
action with Sp1. We speculate that in the TATA-less AR gene promoter,
multiple weak Spl sites at the pur/pyr region adjacent to the GC box c
an provide a readily available source of this transcription factor to
the functional GC box, thereby facilitating the assembly of the initia
tion complex.