| Research Abstract |
To understand the molecular mechanism of action of atrial natriuretic peptide (ANP), the receptors mediating the physiological actions of ANP have been isolated, characterized, classified into subtypes, and visualized at the cellular level. The gene structure of the type C ANP receptor was also determined. 1) Purification and Properties : type C ANP receptor was purified from bovine lung plasma membranes by affinity chromatography and shown to be a disulfide-linked homodimer consisting of two 70-kDa subunits. Amino acid sequences of tryptic fragments of the purified protein were determined. 2) Biosynthetic process of the type C ANP receptor was examined using a cell-free translation system ; glycopeptidase digestion revealed that the mature receptor has three N -linked oligosaccharides per 70-kDa subunit. The ligand-binding activity of the receptor was not affected by the deglycosylation. 3) Immunohistochemical localization : in the kidney and lung, the receptor was localized to the po
… More
docytes of the glomeruli and the epithelial cells, respectively. The glomerular localization explains well the stimulatory effect of ANP on glomerular filtration rate. 4) Mechanism of action : activation by ANP of particulate guanylate cyclase was analyzed by using a specific antireceptor antiserum and by radiation inactivation, and a new dissociative activation mechanism has been proposed. 5) Tissue-specific distribution of AN receptor subtypes was revealed by biochemical, immunochemical, and phrmacological characterization of the receptors on various target tissues. Surprisingly, similar analysis using cultured vascular cells revealed that arterial cells begin to express different subtypes when cultured in vitro and this interesting phenomenon was termed "subtype switching". 6) Subunit structure of the type A ANP receptor : the single chain receptor subtype, termed type A receptor, was shown to be present as a tetramer in the bovine adreal zona glomerulosa cells. 7) The structure of the bovine type C ANP receptor qe was determined and compared with that of the rat type A receptor. The type C receptor gene was unusually long, spanning more than 85 kb. Interesting features are that the genomic structure (exon-intron organization) mimics the domain structure of the receptor and intron positioning is very similar bype A and C receptor genes. Less
|
