Immunochemical studies on the putative plasmalemmal receptor for 1,25-dihydroxyvitamin D3 II. Chick kidney and brain.

Abstract:

:Chick kidney and brain were analyzed for the subcellular distribution (if any) of a putative plasma membrane receptor for 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]. Fractionation protocols were found to be based not only on differential centrifugation conditions, but also gentleness of resuspension procedures, and sufficiently dense Percoll gradients. The postnuclear pellets were resolved on 21.85% Percoll gradients overlayed on 2.4 M sucrose cushions. For both kidney and brain, fraction 1 (bottom of tube) was found to be enriched over whole homogenate 5.4- and 1.6-fold, respectively, in acid phosphatase activity, fractions 2 through 5 were enriched four- and eightfold, respectively, in succinate dehydrogenase activity, fraction 8 contained Golgi, as judged by a small peak of alpha-mannosidase activity, and fraction 9 was enriched sevenfold (for each tissue) in Na+,K+-ATPase activity. Western analyses, using a characterized antibody to the putative chick intestinal plasma membrane vitamin D receptor, revealed the highest levels of antigenicity in both chick kidney and brain in plasma membrane and Golgi fractions, followed by unidentified membranes in fractions 6 and 7 of Percoll gradients. Distribution of specific binding of [3H]1,25(OH)2D3 in Percoll gradient fractions paralleled that of antigenicity. Qualitatively, kidney plasma membrane contained more antigen than brain plasma membrane after Western blot analyses; these results were mirrored by differences in specific binding of the tritiated secosteroid (65 +/- 14.5 and 34 +/- 11.9 fmol/mg of protein, respectively).

journal_name

Steroids

journal_title

Steroids

authors

Jia Z,Nemere I

doi

10.1016/s0039-128x(99)00030-6

subject

Has Abstract

pub_date

1999-08-01 00:00:00

pages

541-50

issue

8

eissn

0039-128X

issn

1878-5867

pii

S0039-128X(99)00030-6

journal_volume

64

pub_type

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