Endocrinology, Vol 119, 2058-2065, Copyright © 1986 by Endocrine Society

ARTICLES

Active calcium transport by porcine thyroid microsomes

Y Nakamura, T Miyamoto, M Koono and S Ohtaki

Microsomal fractions prepared from porcine thyroid glands by differential centrifugations and sucrose density gradient centrifugation showed an ATP-dependent Ca2+ uptake. Electron microscopy and the study of marker enzyme activities suggested that the fractions consisted mainly of endoplasmic reticulum. The amount of transported Ca2+ increased four times in the presence of 20 mM oxalate owing to the precipitation of calcium oxalate, which was detected inside the microsomal vesicles by electron microscopy. Ca2+ was released rapidly when the calcium ionophore, A-23187, was added. The Ca2+ concentration for the half-maximal activation of Ca2+ transport was about 1 microM. These results indicate that Ca2+ is translocated into the lumen of microsomes against a concentration gradient in a manner of the active transport. The microsomes showed Ca2+-dependent ATPase activity and were phosphorylated by the reaction with [gamma-32P]ATP in a similar Ca2+ dependence to that of transport rate. A 105-kDa phosphoprotein was identified by dodecyl sulfate polyacrylamide gel electrophoresis and was found to be sensitive to hydroxylamine. These properties of the phosphoprotein were the same as those of Ca2+ pump ATPase in the endoplasmic reticulum of other cells. These results suggest that the cytosolic Ca2+ is maintained at low levels by the microsomal uptake of Ca2+ by the action of the ATP-dependent Ca2+ pump or active transport system.