CELL SURFACE DNA - AN ANALYSIS OF SPECIFIC RECEPTORS

The proposed study is based on the following observations from our previous
work: (i) Certain peripheral white blood cells possess cell surface DNA
(csDNA), based on the binding of a radioactive DNA probe - viz
lactoferrin. (ii) When cells are stripped of their csDNA (by DNAse) they
will take up exogenous DNA (3H labelled) in a saturable manner consistent
with a ligand receptor relationship - for monocytes and neutrophils the Kd
is about 2.5 x 10-billion M and some 1 to 2.5 x 1 million molecules of
lambda phage DNA bind per cell. (iii) Using a biotin labelled DNA probe we
have demonstrated a DNA binding protein in an SDS-PAGE preparation of
neutrophil surface membranes; the MW of this putative DNA receptor was
about 30,000. (iv) Preliminary studies of 3H DNA binding in normals and
patients with connective tissue diseases indicates that some patients with
systemic lupus erythematous (SLE) have an apparent DNA-receptor defect. It is intended to expand these findings as follows: (i) Establish which
cell populations are capable of binding DNA; (ii) Verify that the binding
is a saturable phenomenon with a Kd consistent with a ligand-receptor
association; (iii) Determine whether bound DNA is internalized, and if so,
if it is further degraded; (iv) Study factors influencing receptor turnover
and regeneration; (v) Analyze purified membrane preparations from different
cell types (using the biotin labelled DNA probe/PAGE system) for DNA
binding proteins; (vi) Study the prevalence of deficient DNA binding in
patients with SLE and see if this correlates with disease activity,
serological markers inherited complement deficiencies or HLA typing; (vii)
Look for abnormal DNA binding in other connective tissue disorders, other
chronic diseases and normals; (viii) Try to determine whether deficient DNA
binding is due to a receptor defect or a serum blocking factor. The definitive demonstration of a DNA receptor on certain immunocompetent
cells would have important implications for molecular biology in general,
and the confirmation of a receptor defect in SLE would open up a new avenue
of research into the pathogenesis of this prototypical autoimmune disease.