Neuronal nitric oxide synthase interacts with Sp1 through the PDZ domain inhibiting Sp1-mediated copper-zinc superoxide dismutase expression.
In this report we demonstrate that neuronal nitric oxide synthase (nNOS) is able to interact with Sp1 both in vivo and in vitro. In particular, we show that such interaction is mediated by the N-terminal PDZ domain of full length nNOS (fl-nNOS). In fact nNOS mutant lacking the PDZ domain (ΔnNOS) displays an impaired ability to bind to Sp1, as demonstrated by co-immunoprecipitation experiments. The overexpression of fl-nNOS in SH-SY5Y cells leads to the formation of nNOS/Sp1 heterocomplex and inhibits the binding of Sp1 to DNA. Among the Sp1 target genes we looked at the possible alteration of binding to copper-zinc superoxide dismutase gene (sod1) promoter. We find that the interaction of nNOS with Sp1 leads to a significant decrease of SOD1 mRNA, protein level and activity. The overexpression of ΔnNOS results in an inability to sequester Sp1 and unaffected Sp1 DNA binding capacity, allowing sod1 to be expressed. The data reported give effort to the possible involvement of nNOS in regulating gene transcription in NO-independent manner giving an additional significance to the expression of specific nNOS splicing variants.
Dept. of Biology, University of Rome "Tor Vergata", Via della Ricerca Scientifica 1, 00133 Rome, Italy.
Sara
S
Baldelli
Katia
K
Aquilano
Giuseppe
G
Rotilio
Maria R
MR
Ciriolo
D000818:Animals; D003300:Copper / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D006801:Humans; D007136:Immunoglobulins / Q000235:genetics / Q000378:metabolism; D047468:Immunoprecipitation; D051379:Mice; D009474:Neurons / Q000378:metabolism; D009569:Nitric Oxide / Q000032:analysis / Q000378:metabolism; D052248:Nitric Oxide Synthase Type I / Q000235:genetics / Q000378:metabolism; D054731:PDZ Domains / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D051381:Rats; D013482:Superoxide Dismutase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D014407:Tumor Cells, Cultured; D015032:Zinc / Q000378:metabolism
D007136:Immunoglobulins; D012333:RNA, Messenger; C090698:SP1 antigen; D009569:Nitric Oxide; D003300:Copper; D052248:Nitric Oxide Synthase Type I; D013482:Superoxide Dismutase; D015032:Zinc