Direct binding of atrial natriuretic factor to adrenocortical mitochondria.
Atrial natriuretic factor (ANF) is a known antagonist of adrenocortical aldosterone synthesis and secretion. Immunocytochemical and ultrastructural autoradiographic evidence suggests that ANF may bind to mitochondria of a number of target tissues including adrenal cortex. Consequently, the ability of [125I]ANF to bind directly to isolated bovine adrenocortical mitochondria was assessed. Mitochondrial-enriched subfractions of adrenocortical homogenates were prepared by differential and sucrose gradient centrifugation. Mitochondrial membranes specifically bound [125I]ANF. At 20 degrees C equilibrium was achieved between 90 and 120 min. [125I]ANF binding was inhibited in a concentration-dependent manner by unlabelled ANF (IC50 about 5 x 10(-10) M); other substances with biological actions on glomerulosa cells (arginine vasopressin, angiotensin II) did not alter [125I]ANF binding. Similarly shorter ANF fragments including ANF-(103-125), ANF-(99-109) and ANF-(111-126) had no significant competitive effect on binding of the labelled ligand. While Ca2+ and Mg2+ had little effect on ANF binding, the divalent cation Ni2+ inhibited binding of radiolabelled ANF by 90% (IC50 about 8.3 x 10(-5) M). Scatchard analysis revealed both high and low affinity binding sites for [125I]ANF with respective KDs of 4.7 +/- 7 pM and 0.3 +/- 0.02 nM and receptor densities of 1.1 +/- 0.2 and 8.6 +/- 0.1 pmol/mg protein. At 0.2 nM, Ni2+ caused a 5-fold and 100-fold decrease in high and low affinity [125I]ANF binding, respectively. The data demonstrate that ANF binds directly to mitochondria and perhaps it is at this site that the atrial peptide negatively modulates agonist-induced aldosterone biosynthesis.
Heisler
S
S
Unité de Biorégulation cellulaire, Le Centre hospitalier de l'Université Laval, Sainte-Foy Québec, Canada.
D000302:Adrenal Cortex / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D015415:Biomarkers; D002417:Cattle; D066298:In Vitro Techniques; D007457:Iodine Radioisotopes; D008297:Male; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D009532:Nickel / Q000494:pharmacology; D051381:Rats; D013347:Subcellular Fractions / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D013997:Time Factors
D015415:Biomarkers; D007457:Iodine Radioisotopes; D009532:Nickel; D009320:Atrial Natriuretic Factor