Metabolic modulation of hexokinase association with mitochondria in living smooth muscle cells. Lynch, Ronald M., Walter Carrington, Kevin E. Fogarty, and Fredric S. Fay. Departments of Physiology and Pharmacology, University of Arizona, Health Sciences Center, Tucson AZ 85724, Tel. (602) 626-2472, FAX (602) 626-2383 and Department of Physiology, and Program in Molecular Medicine. University of Massachusetts Medical Center, Worcester MA 01655, Tel. (508) 856-2346, FAX (508) 856-1840.
APStracts 2:0309C, 1995.
Hexokinase isoform I binds to mitochondria of many cell types. It has been hypothesized that this association is regulated by changes in the concentrations of specific cellular metabolites. To study the distribution of hexokinase in living cells, fluorophore labeled functional hexokinase I was prepared. Following microinjection into A7R5 smooth muscle cells, hexokinase localized to distinct structures identified as mitochondria. The endogenous hexokinase demonstrated a similar distribution using immunocytochemistry. 2-deoxyglucose elicited an increase in glucose-6-phosphate and a decrease in ATP levels, and diminished hexokinase binding to mitochondria in single cells. 3-O-methyl glucose elicited slow developing decreases in all three parameters. By contrast, cyanide elicited a rapid decrease in both ATP and hexokinase binding. Analysis of changes in metabolite levels and hexokinase binding indicate a positive correlation between binding and cell energy state as monitored by ATP. On the other hand, only in the presence of 2-deoxyglucose was the predicted inverse correlation between binding and G-6-P observed. Unlike the relatively large changes in distribution observed with the fluorescent-injected hexokinase, cyanide caused only a small decrease in the localization of endogenous hexokinase with mitochondria. These findings suggest that changes in the concentrations of specific metabolites can alter the binding of hexokinase I to specific sites on mitochondria. Moreover, the apparent difference in sensitivity of injected and endogenous hexokinase to changes in metabolites may reflect the presence of at least two classes of binding mechanisms for hexokinase, with differential sensitivity to metabolites. 

Received 21 November 1994; accepted in final form 21 August 1995.
APS Manuscript Number C677-4.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 15 September 1995.