Jp. Quinn et al., MULTIPLE PROTEIN COMPLEXES, INCLUDING AP2 AND SP1, INTERACT WITH A SPECIFIC SITE WITHIN THE RAT PREPROTACHYKININ-A PROMOTER, Biochimica et biophysica acta, N. Gene structure and expression, 1263(1), 1995, pp. 25-34
We demonstrate that there is a unique AP2 binding site in the rat prep
rotachykinin-A promoter (rPPT) spanning - 865 to - 47. AP2 is a transc
ription factor whose expression in sensory neurons has been correlated
with rPPT expression in these cells. This binding site is adjacent to
an element we previously identified as binding a single stranded DNA
binding protein which was also present in sensory neurons. These two c
omplexes encompass a region which we had proposed might form a stem-lo
op structure, allowing binding of the single stranded DNA binding prot
ein to the DNA. Here using electrophoretic mobility shift analysis we
demonstrate that the DNA region corresponding to the putative stem-loo
p structure is bound by a variety of transcription factors, including
in addition to AP2 the ubiquitous Spl. DNase 1 footprint analysis demo
nstrates that binding to this domain by the proteins recognising the d
ouble-stranded form of the cis acting element is mutually exclusive. A
promoter fragment containing this domain demonstrated a DNase 1 footp
rint over the 5' region of the stem-loop structure. Competition of the
binding for this element by an oligonucleotide corresponding to the s
tem-loop structure removed the 5' footprint and exposed a new footprin
t over the 3' region of the stem-loop structure and extending for seve
ral base pairs. This change in protection observed with DNase 1 digest
ion also correlated with changes of the DNase 1 pattern at specific lo
cations 3' of the proposed stem-loop structure. These changes correlat
ed with two DNA sequences which were homologous to one another and to
a region within the proposed stem-loop structure. Our results indicate
that AP2 could regulate rPPT gene expression by a variety of mechanis
ms.