We have used quantitative epifluorescence microscopy of fluorescent ATP to measure single-nucleotide turnover in skinned skeletal muscle fibers from mouse models of female aging and hormone treatment. Aging causes declines in muscle strength, often leading to frailty, disability, and loss of independence for the elderly. Female muscle is additionally affected by age due to reduction of ovarian hormone production with menopause. Estradiol (E2) is the key hormonal signal to skeletal muscle in females, and strength loss is attenuated by E2 treatment. To investigate E2 mechanisms on skeletal muscle, single fibers were isolated from sham-operated or ovariectomized (OVX) mice, with or without E2 treatment, and were incubated with 2′-(or-3′)-O-(N-methylanthraniloyl) adenosine 5′-triphosphate (mantATP). We measured decay of mantATP fluorescence in an ATP-chase experiment, as pioneered by Cooke and coworkers, who unveiled a novel regulated state of muscle myosin characterized by slow nucleotide turnover on the order of minutes, termed the super-relaxed state (SRX). We detected a slow phase of nucleotide turnover in a portion of the myosin heads from sham fibers, consistent with SRX. Turnover was substantially faster in OVX fibers, with a turnover time constant for the slow phase of 65 ± 8 s as compared to 102 ± 7 s for sham fibers. 60-days E2 treatment in OVX mice substantially reversed this effect on SRX, while acute exposure of isolated muscles from OVX mice to E2 had no effect. We conclude that E2-mediated signaling reversibly regulates slow ATP turnover by myosin. Age- and hormone-related muscle functional losses may be targetable at the level of myosin structure/function for strategies to offset weakness and metabolic changes that occur with age.
|Original language||English (US)|
|Number of pages||5|
|Journal||Biochemical and Biophysical Research Communications|
|State||Published - Jan 2 2015|
Bibliographical noteFunding Information:
We thank Simon Gruber for helpful discussions and Octavian Cornea for helping to prepare the manuscript. We thank Roger Cooke for providing expert advice about super-relaxation. Fluorescence microscopy was performed at the University of Minnesota Imaging Centers with assistance from Guillermo Marqués. This work was supported primarily by NIH grants to DDT ( R37 AG26160 and R37 AG26160-09S2 ). Support was also provided by NIH grants to DAL (R01 AG31743) and BAC (K99 HL122397). KJP and BCC were supported by NIH grant T32 AR7612 .
© 2014 Elsevier Inc. All rights reserved.
- Nucleotide turnover
- Regulatory light chain
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