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Activity-dependent and -independent nuclear fluxes of HDAC4 mediated by different kinases in adult skeletal muscle.

Liu Y, Randall WR, Schneider MF - J. Cell Biol. (2005)

Bottom Line: Class II histone deacetylases (HDACs) may decrease slow muscle fiber gene expression by repressing myogenic transcription factor myocyte enhancer factor 2 (MEF2).Thus, calcium transients for slow, but not fast, fiber stimulation patterns appear to provide sufficient Ca(2+)-dependent activation of nuclear CaMKII to result in net nuclear efflux of HDAC4.Nucleocytoplasmic shuttling of HDAC4-GFP in unstimulated resting fibers was not altered by KN-62, but was blocked by staurosporine, indicating that different kinases underlie nuclear efflux of HDAC4 in resting and stimulated muscle fibers.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, MD 21201, USA.

ABSTRACT
Class II histone deacetylases (HDACs) may decrease slow muscle fiber gene expression by repressing myogenic transcription factor myocyte enhancer factor 2 (MEF2). Here, we show that repetitive slow fiber type electrical stimulation, but not fast fiber type stimulation, caused HDAC4-GFP, but not HDAC5-GFP, to translocate from the nucleus to the cytoplasm in cultured adult skeletal muscle fibers. HDAC4-GFP translocation was blocked by calmodulin-dependent protein kinase (CaMK) inhibitor KN-62. Slow fiber type stimulation increased MEF2 transcriptional activity, nuclear Ca(2+) concentration, and nuclear levels of activated CaMKII, but not total nuclear CaMKII or CaM-YFP. Thus, calcium transients for slow, but not fast, fiber stimulation patterns appear to provide sufficient Ca(2+)-dependent activation of nuclear CaMKII to result in net nuclear efflux of HDAC4. Nucleocytoplasmic shuttling of HDAC4-GFP in unstimulated resting fibers was not altered by KN-62, but was blocked by staurosporine, indicating that different kinases underlie nuclear efflux of HDAC4 in resting and stimulated muscle fibers.

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Time course of nuclear to cytoplasmic translocation of HDAC4 during 10-Hz train stimulation. (A) The average fluorescence intensity per pixel over whole nuclei (closed circles) or over the cytoplasm (open circles) was quantitated and normalized as described in Materials and methods. 5-s trains of 10-Hz stimuli every 50 s resulted in net nuclear to cytoplasmic translocation of HDAC4-GFP. Nuclear fluorescence declined continuously during the 120-min stimulation period. The cytoplasmic fluorescence remained constant during the same period of time. (B) The reversibility of HDAC4-GFP efflux due to electrical stimulation. (C) CaMK inhibitor KN-62 blocked the nuclear to cytoplasmic translocation of HDAC4-GFP in stimulated fibers.
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fig2: Time course of nuclear to cytoplasmic translocation of HDAC4 during 10-Hz train stimulation. (A) The average fluorescence intensity per pixel over whole nuclei (closed circles) or over the cytoplasm (open circles) was quantitated and normalized as described in Materials and methods. 5-s trains of 10-Hz stimuli every 50 s resulted in net nuclear to cytoplasmic translocation of HDAC4-GFP. Nuclear fluorescence declined continuously during the 120-min stimulation period. The cytoplasmic fluorescence remained constant during the same period of time. (B) The reversibility of HDAC4-GFP efflux due to electrical stimulation. (C) CaMK inhibitor KN-62 blocked the nuclear to cytoplasmic translocation of HDAC4-GFP in stimulated fibers.

Mentions: Next, we investigated translocation of HDAC4-GFP from the nucleus to the cytoplasm in response to electrical stimulation patterns mimicking the physiological activity patterns of skeletal muscle. After 30 min without stimulation, during which the fluorescence in both the nucleus and the cytoplasm was stable (Fig. 1, −30 and 0 min), fibers were repeatedly stimulated with a 10-Hz train for 5 s every 50 s. Field stimulation resulted in visible twitches throughout the period of stimulation in all fibers used for analysis. This electrical stimulation caused a noticeable translocation of HDAC4-GFP from the nucleus to the cytoplasm (Fig. 1, 60 and 120 min). Over a 2-h period of repetitive stimulation (5-s duration 10-Hz trains every 50 s) nuclear fluorescence continuously decreased, indicating continued net translocation of HDAC4-GFP out of the nucleus. This net efflux of HDAC4-GFP from the nucleus occurred without any significant change in cytoplasmic fluorescence (Fig. 2 A), indicating that the total cytoplasmic pool of HDAC4-GFP was much larger than the size of the total nuclear pool.


Activity-dependent and -independent nuclear fluxes of HDAC4 mediated by different kinases in adult skeletal muscle.

Liu Y, Randall WR, Schneider MF - J. Cell Biol. (2005)

Time course of nuclear to cytoplasmic translocation of HDAC4 during 10-Hz train stimulation. (A) The average fluorescence intensity per pixel over whole nuclei (closed circles) or over the cytoplasm (open circles) was quantitated and normalized as described in Materials and methods. 5-s trains of 10-Hz stimuli every 50 s resulted in net nuclear to cytoplasmic translocation of HDAC4-GFP. Nuclear fluorescence declined continuously during the 120-min stimulation period. The cytoplasmic fluorescence remained constant during the same period of time. (B) The reversibility of HDAC4-GFP efflux due to electrical stimulation. (C) CaMK inhibitor KN-62 blocked the nuclear to cytoplasmic translocation of HDAC4-GFP in stimulated fibers.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171787&req=5

fig2: Time course of nuclear to cytoplasmic translocation of HDAC4 during 10-Hz train stimulation. (A) The average fluorescence intensity per pixel over whole nuclei (closed circles) or over the cytoplasm (open circles) was quantitated and normalized as described in Materials and methods. 5-s trains of 10-Hz stimuli every 50 s resulted in net nuclear to cytoplasmic translocation of HDAC4-GFP. Nuclear fluorescence declined continuously during the 120-min stimulation period. The cytoplasmic fluorescence remained constant during the same period of time. (B) The reversibility of HDAC4-GFP efflux due to electrical stimulation. (C) CaMK inhibitor KN-62 blocked the nuclear to cytoplasmic translocation of HDAC4-GFP in stimulated fibers.
Mentions: Next, we investigated translocation of HDAC4-GFP from the nucleus to the cytoplasm in response to electrical stimulation patterns mimicking the physiological activity patterns of skeletal muscle. After 30 min without stimulation, during which the fluorescence in both the nucleus and the cytoplasm was stable (Fig. 1, −30 and 0 min), fibers were repeatedly stimulated with a 10-Hz train for 5 s every 50 s. Field stimulation resulted in visible twitches throughout the period of stimulation in all fibers used for analysis. This electrical stimulation caused a noticeable translocation of HDAC4-GFP from the nucleus to the cytoplasm (Fig. 1, 60 and 120 min). Over a 2-h period of repetitive stimulation (5-s duration 10-Hz trains every 50 s) nuclear fluorescence continuously decreased, indicating continued net translocation of HDAC4-GFP out of the nucleus. This net efflux of HDAC4-GFP from the nucleus occurred without any significant change in cytoplasmic fluorescence (Fig. 2 A), indicating that the total cytoplasmic pool of HDAC4-GFP was much larger than the size of the total nuclear pool.

Bottom Line: Class II histone deacetylases (HDACs) may decrease slow muscle fiber gene expression by repressing myogenic transcription factor myocyte enhancer factor 2 (MEF2).Thus, calcium transients for slow, but not fast, fiber stimulation patterns appear to provide sufficient Ca(2+)-dependent activation of nuclear CaMKII to result in net nuclear efflux of HDAC4.Nucleocytoplasmic shuttling of HDAC4-GFP in unstimulated resting fibers was not altered by KN-62, but was blocked by staurosporine, indicating that different kinases underlie nuclear efflux of HDAC4 in resting and stimulated muscle fibers.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, MD 21201, USA.

ABSTRACT
Class II histone deacetylases (HDACs) may decrease slow muscle fiber gene expression by repressing myogenic transcription factor myocyte enhancer factor 2 (MEF2). Here, we show that repetitive slow fiber type electrical stimulation, but not fast fiber type stimulation, caused HDAC4-GFP, but not HDAC5-GFP, to translocate from the nucleus to the cytoplasm in cultured adult skeletal muscle fibers. HDAC4-GFP translocation was blocked by calmodulin-dependent protein kinase (CaMK) inhibitor KN-62. Slow fiber type stimulation increased MEF2 transcriptional activity, nuclear Ca(2+) concentration, and nuclear levels of activated CaMKII, but not total nuclear CaMKII or CaM-YFP. Thus, calcium transients for slow, but not fast, fiber stimulation patterns appear to provide sufficient Ca(2+)-dependent activation of nuclear CaMKII to result in net nuclear efflux of HDAC4. Nucleocytoplasmic shuttling of HDAC4-GFP in unstimulated resting fibers was not altered by KN-62, but was blocked by staurosporine, indicating that different kinases underlie nuclear efflux of HDAC4 in resting and stimulated muscle fibers.

Show MeSH
Related in: MedlinePlus