Z. Khaled et al., MULTIPLE MECHANISMS MAY CONTRIBUTE TO THE CELLULAR ANTIADHESIVE EFFECTS OF PHOSPHOROTHIOATE OLIGODEOXYNUCLEOTIDES, Nucleic acids research, 24(4), 1996, pp. 737-745
Phosphorothioate oligodeoxynucleotides complementary to the p65 (Rel A
) subunit of the NF-kappa B nuclear transcriptional regulatory factor
have been suggested to be sequence specific blockers of cellular adhes
ion. We studied the effects of Rel A antisense, Rel A sense and other
phosphorothioate oligodeoxynucleotides on cellular adhesion and found
that blockade of adhesion was predominately non-sequence specific, Pho
sphorothioate oligodeoxynucleotides bind to the extracellular matrix (
ECM) of NIH 3T3 cells, and to the ECM elements laminin and fibronectin
. By use of a gel mobility shift assay, the association of the A subun
it of laminin with a probe 12mer phosphodiester oligodeoxynucleotide c
ould be demonstrated. This interaction was described by a single-site
binding equation (K-d = 14 mu M). Human Rel A antisense and sense olig
odeoxynucleotides, and two synthetic persulfated heparin analogs were
excellent competitors of the binding of the probe oligodeoxynucleotide
to laminin. Taken together, these data indicate that oligodeoxynucleo
tide binding occurred at or near the heparin-binding site. Competition
for 5' P-32-SdT18 (an 18mer phosphorothioate homopolymer of thymidine
) binding to fibronectin with the discrete heparin analogs, as well as
with SdC(28), was also observed. Phosphorothioate oligodeoxynucleotid
es (Rel A antisense >> Rel A sense) inhibited the binding of laminin t
o bovine brain sulfatide, but not to its cell surface receptors on MCF
-7 cells. By flow cytometric analysis we have also shown, in contrast
to what was observed with laminin, that phosphorothioates non-specific
ally block the specific binding of fluoresceinated fibronectin to its
cell surface receptors on phorbol-12,13-myristate acetate-treated Jurk
at cells. Blockade of specific binding occurred in the oligodeoxynucle
otide treated cells in the presence or absence of oligomer in the medi
a.