Characterization of the AT(4) receptor in a human neuroblastoma cell line (SK-N-MC).
Angiotensin IV (Ang IV), the 3-8 fragment of angiotensin II (Ang II), binds to a distinct receptor designated the AT(4) receptor. The peptide elicits a range of vascular and central actions including facilitation of memory retention and retrieval in several learning paradigms. The aim of this study was to characterize the AT(4) receptor in a human cell line of neural origin. Receptor binding studies indicate that the human neuroblastoma cell line SK-N-MC cells express a high-affinity Ang IV binding site with a pharmacological profile similar to the AT(4) receptor: (125)I]-Ang IV and (125)I]-Nle(1)-Ang IV bind specifically to the SK-N-MC cell membranes (K(d) = 0.6 and 0.1 nM) in a saturable manner (B(max) = 1.2 pmol/mg of protein). AT(4) receptor ligands, Nle(1)-Ang IV, Ang IV and LVV-haemorphin 7 (LVV-H7), compete for the binding of [(125)I]-Ang IV or [(125)I]-Nle(1)-Ang IV to the SK-N-MC cell membranes with rank order potencies of Nle(1)-Ang IV > Ang IV > LVV-H7 with IC(50) values of 1.4, 8.7 and 59 nM ([(125)I]-Ang IV) and 1.8, 20 and 168 nM ([(125)I]-Nle(1)-Ang IV), respectively. The binding of [(125)I]-Ang IV or [(125)I]-Nle(1)-Ang IV to SK-N-MC cell membranes was not affected by the presence of GTP gamma S. Both Ang IV and LVV-H7 stimulated DNA synthesis in this cell line up to 72 and 81% above control levels, respectively. The AT(4) receptor in the SK-N-MC cells is a 180-kDa glycoprotein; under non-reducing conditions a 250-kDa band was also observed. In summary, the human neuroblastoma cell line, SK-N-MC, expresses functional AT(4) receptors that are responsive to Ang IV and LVV-H7, as indicated by an increase in DNA synthesis. This is the first human cell line of neural origin shown to express the AT(4) receptor.
Mustafa
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Howard Florey Institute of Experimental Physiology and Medicine, University of Melbourne, Parkville, Australia.
Chai
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Mendelsohn
F A
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Moeller
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Albiston
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D000804:Angiotensin II / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000494:pharmacology; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D006031:Glycosylation; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006454:Hemoglobins / Q000494:pharmacology; D006801:Humans; D007457:Iodine Radioisotopes; D007700:Kinetics; D008024:Ligands; D009447:Neuroblastoma; D010446:Peptide Fragments / Q000494:pharmacology; D011869:Radioligand Assay; D011945:Receptors, Angiotensin / Q000187:drug effects / Q000378:metabolism*; D013936:Thymidine / Q000378:metabolism; D014407:Tumor Cells, Cultured
C090830:AT4 receptor; D003432:Cross-Linking Reagents; D006454:Hemoglobins; D007457:Iodine Radioisotopes; D008024:Ligands; D010446:Peptide Fragments; D011945:Receptors, Angiotensin; D000804:Angiotensin II; C012877:angiotensin II, des-Asp(1)-des-Arg(2)-Ile(5)-; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C074309:LVV-hemorphin-7; D013936:Thymidine