Cloning and expression of angiotensin II type 2 (AT2) receptors from murine neuroblastoma N1E-115 cells: evidence for AT2 receptor heterogeneity.
Homology-based PCR was used to isolate angiotensin II type 2 (AT2) receptor cDNA from murine neuroblastoma N1E-115 cells. Despite subtle differences in the nucleotide sequence (the N1E-115 clone coded for Phe133 as TTC and Gln326 as CAG; base substitutions are in bold-italics), the AT2 receptor protein was identical to other reported murine AT2 clones. When transfected into COS-1 cells, the expressed AT2 receptor displayed high affinity for AngII and for AT2-selective compounds, GTP gamma S-insensitive agonist binding and enhanced agonist binding by dithiothreitol. Previously, we have demonstrated that N1E-115 cells possess two distinct subpopulations of AT2 receptors, defined as peak I and peak III receptors, that can be separated by heparin-sepharose chromatography. The two subpopulations differ pharmacologically, biochemically and immunologically. The binding properties of the cloned AT2 receptor closely resembled that of peak III receptors. Moreover, antisera raised against peak I AT2 receptors failed to immunoreact to either peak III receptors or cloned AT2 receptors expressed in COS-1 cells. Collectively, these data suggest that the cloned AT2 receptor is identical to peak III receptors from N1E-115 cells and that a novel AT2 receptor (peak I) remains to be cloned.
Yee
D K
DK
Department of Animal Biology, University of Pennsylvania, Philadelphia 19104, USA.
He
P
P
Yang
X D
XD
Reagan
L P
LP
Hines
J
J
Siemens
I R
IR
Fluharty
S J
SJ
D000804:Angiotensin II / Q000378:metabolism / Q000494:pharmacology; D057911:Angiotensin Receptor Antagonists; D000818:Animals; D001713:Biphenyl Compounds / Q000494:pharmacology; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004229:Dithiothreitol / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D008125:Locus Coeruleus / Q000378:metabolism; D019808:Losartan; D051379:Mice; D009447:Neuroblastoma; D009842:Oligopeptides / Q000494:pharmacology; D009847:Olivary Nucleus / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011725:Pyridines / Q000494:pharmacology; D044139:Receptor, Angiotensin, Type 2; D011945:Receptors, Angiotensin / Q000096:biosynthesis* / Q000502:physiology; D011994:Recombinant Proteins / Q000096:biosynthesis; D013777:Tetrazoles / Q000494:pharmacology; D014162:Transfection; D014407:Tumor Cells, Cultured
D057911:Angiotensin Receptor Antagonists; D001713:Biphenyl Compounds; D018076:DNA, Complementary; D007093:Imidazoles; D009842:Oligopeptides; D011725:Pyridines; D044139:Receptor, Angiotensin, Type 2; D011945:Receptors, Angiotensin; D011994:Recombinant Proteins; D013777:Tetrazoles; D000804:Angiotensin II; C060894:CGP 42112A; C073402:PD 123319; D019808:Losartan; D004229:Dithiothreitol