Microtubules Mediate Cellular 25-Hydroxyvitamin D3 Trafficking and the Genomic Response to 1,25-Dihydroxyvitamin D3 in Normal Human Monocytes

doi:10.1074/jbc.270.38.22160

Microtubules Mediate Cellular 25-Hydroxyvitamin D₃ Trafficking and the Genomic Response to 1,25-Dihydroxyvitamin D₃ in Normal Human Monocytes (*)

From the Department of Internal Medicine, Renal Division, and the
Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110-1093

^¶ To whom correspondence and reprint requests should be addressed:
Dept. of Internal Medicine, Renal Division, Washington University School of Medicine, 660 South Euclid Ave., Box 8126, St. Louis, MO 63110-1093.
Tel.: 314-362-8248; Fax: 314-362-8237.

Abstract

The genomic actions of 1,25-dihydroxyvitamin D (1,25(OH)D) are mediated by the intracellular vitamin D receptor (VDR). Although immunocytochemistry has shown that disruption of microtubular assembly prevents nuclear access of the sterol-VDR complex, the role of microtubules in the response to 1,25(OH)D has not been studied in viable cells. Our studies examined this interaction in normal human monocytes. Monocytes convert 25(OH)D to 1,25(OH)D and to 24-hydroxylated metabolites more polar than 1,25(OH)D. Microtubule disruption totally abolished the ability of exogenous 1,25(OH)D to suppress its own synthesis and to induce 24-hydroxylase mRNA and activity, without affecting either total 1,25(OH)D uptake or maximal 1,25(OH)D-VDR binding. Thus, intact microtubules are essential for 1,25(OH)D-dependent modulation of gene transcription. Interestingly, microtubule disruption also decreased monocyte 1,25(OH)D synthesis, not by decreasing the V_max of monocyte mitochondrial 1α-hydroxylase but through an increase in the K for 25(OH)D. We examined 25(OH)D transport. Microtubule disruption did not affect total cellular 25(OH)D uptake but reduced its intracellular trafficking to the mitochondria. Thus, microtubules participate in intracellular 25(OH)D transport, and their integrity determines normal 1,25(OH)D synthesis.

Footnotes

↵^§ Recipient of a research fellowship from Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Argentina.
↵* This work was supported in part by United States Public Health Service NIDDK, National Institutes of Health Grants DK-09976 and DK-07126. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

1,25(OH)D

1,25-dihydroxyvitamin D

VDR

vitamin D receptor; 25(OH)D, 25-hydroxyvitamin D

HPLC

high performance liquid chromatography

PBS

phosphate-buffered saline

PIPES

piperazine-N,N‘-bis(2-ethanesulfonic acid) or 1,4-piperazinediethanesulfonic acid.
↵²S. Kamimura, M. Gallieni, E. Slatopolsky, and A. Dusso, unpublished data.
- Received April 21, 1995.