D. Lang et T. Stamminger, MINOR-GROOVE CONTACTS ARE ESSENTIAL FOR AN INTERACTION OF THE HUMAN CYTOMEGALOVIRUS IE2 PROTEIN WITH ITS DNA TARGET, Nucleic acids research, 22(16), 1994, pp. 3331-3338
The 86 kDa immediate early-2 protein (IE2, IE86) of human cytomegalovi
rus (HCMV) is a multifunctional polypeptide that can regulate gene exp
ression both positively and negatively. In particular, it represses it
s own mRNA synthesis by binding directly to a sequence element, termed
cis repression signal (CRS), that is located between the TATA box and
the transcriptional start site of the major IE enhancer/promoter of H
CMV. Here, we provide evidence that IE86, unlike most sequence-specifi
c DNA-binding proteins, interacts primarily within the minor groove of
the DNA helix, This was shown by hydroxyl radical and methylation int
erference assays. In addition, binding studies with inosine-substitute
d oligonucleotides which have an altered major groove morphology witho
ut changing the surface of the minor groove, confirmed the results obt
ained in interference analyses. This establishes IE86 as a member of a
small group of DNA binding proteins that interact with A - T rich seq
uences within the minor groove and which also includes the TATA-box bi
nding protein TBP. Remarkably, IE86 and TBP are able to bind simultane
ously in an immediate vicinity at the major IE enhancer/promoter of HC
MV. As minor groove binding proteins are known to bend DNA heavily thi
s could contribute to the observed negative regulation of transcriptio
n by IE86.