A. Ogo et al., CAMP-DEPENDENT TRANSACTIVATION INVOLVING THE HOMEODOMAIN PROTEIN PBX1, Archives of biochemistry and biophysics, 338(2), 1997, pp. 193-200
Pbx1 is a DNA-binding homeodomain protein originally discovered in the
t(1;19) chromosomal translocation associated with pediatric pre-B acu
te lymphoblastic leukemia. Previously we reported a cAMP-regulatory se
quence (CRS1) in the promoter region of the bovine CYP17 gene encoding
steroid 17 alpha-hydroxylase cytochrome P450 (P450c17) to be the firs
t endogenous Pbx1 binding site and that overexpression of Pbx1 in mous
e adrenal Y1 tumor cells enhances cAMP-dependent transcription mediate
d by this element. Here we report further characterization of Pbx1 bin
ding site in CRS1 and role of Pbx1 in cAMP-dependent, CRS1-mediated tr
anscription. By gel shift analysis utilizing nuclear extracts from Y1
cells, a high-affinity Pbx-binding sequence has been determined to be
TTGAT(T/ G)GA(T/C)A which represents the 5' portion of CRS1. An artifi
cial Pbx-binding sequence (PRS), previously determined by random PCR a
nalysis, is similar to the Pbx1-binding sequence in CRS1 and by both g
el shift analysis and transfection studies shows characteristics very
similar to CRS1. Upon overexpression, Pbx1 is found capable of enhanci
ng CRS1-mediated transcription in both steroidogenic (Y1, JEG3) and no
nsteroidogenic (HepG2 and S194) cells when coexpressed with the cataly
tic subunit of cAMP-dependent protein kinase A. Thus even though Pbx1
has been found to be involved only in cAMP-dependent transcription of
a gene involved in steroidogenesis (CYP17), Pbx1 is capable of partici
pating in cAMP-dependent transcription of target genes without complex
formation with steroidogenic tissue-specific nuclear factors. (C) 199
7 Academic Press.