Limits...
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.

Show MeSH

Related in: MedlinePlus

The subcellular distribution of CaM-YFP was not changed by 10- or 1-Hz electrical stimulation. (A) A typical living fiber expressing CaM-YFP is shown for 30 min (−30 and 0) before stimulation and 60 and 120 min after repetitive stimulation with 10-Hz trains. In resting fibers, CaM-YFP was present in the cytoplasm as well as the nucleus. (B) The fluorescence signals from both the nucleus and the cytoplasm were quantitated. Data are presented as the ratio of the average fluorescence intensity per pixel from the nucleus relative to the cytoplasm. Neither 10-Hz train stimulation (left) for 2 h nor 1-Hz continuous stimulation (right) resulted in any subcellular redistribution of CaM-YFP. (C) FRAP of CaM-YFP was performed in the nuclear area of a muscle fiber. Shown is a nucleus before photobleaching, immediately after photobleaching, and after 30 and 60 min of recovery. The entire nuclear region was selectively bleached. Bars, 10 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2171787&req=5

fig6: The subcellular distribution of CaM-YFP was not changed by 10- or 1-Hz electrical stimulation. (A) A typical living fiber expressing CaM-YFP is shown for 30 min (−30 and 0) before stimulation and 60 and 120 min after repetitive stimulation with 10-Hz trains. In resting fibers, CaM-YFP was present in the cytoplasm as well as the nucleus. (B) The fluorescence signals from both the nucleus and the cytoplasm were quantitated. Data are presented as the ratio of the average fluorescence intensity per pixel from the nucleus relative to the cytoplasm. Neither 10-Hz train stimulation (left) for 2 h nor 1-Hz continuous stimulation (right) resulted in any subcellular redistribution of CaM-YFP. (C) FRAP of CaM-YFP was performed in the nuclear area of a muscle fiber. Shown is a nucleus before photobleaching, immediately after photobleaching, and after 30 and 60 min of recovery. The entire nuclear region was selectively bleached. Bars, 10 μm.

Mentions: Because CaMK appears to mediate the stimulation-dependent translocation of HDAC4 from the nucleus to the cytoplasm (Fig. 2), we next examined the intracellular distribution of CaM-YFP, expressed by adenoviral infection, to see if stimulation recruited CaM to the nucleus. In resting fibers, CaM-YFP exhibited a sarcomeric pattern in the cytoplasm, with the bright lines in the pattern corresponding to α-actinin localization at the Z line (not depicted), and a diffuse pattern in the nucleus except the nucleolus (Fig. 6 A). There was significant CaM-YFP in the A-band region between Z-lines, giving a pixel fluorescence about half that at the Z-line and indicating presence of CaM within the sarcomere away from the Z-line. The mean value of the ratio of nuclear to cytoplasmic mean pixel fluorescence was 0.79 ± 0.05 (11 nuclei from 7 fibers). This ratio was stable in resting fibers and did not change during a 2-h period of repetitive stimulation with 5-s trains of 10-Hz stimuli applied every 50 s (Fig. 6 B, left), a stimulus pattern that produced clear KN-62-sensitive, stimulation-dependent translocation of HDAC4 from the nucleus to the cytoplasm (Fig. 2). CaM-YFP also did not exhibit any change in nuclear-cytoplasmic distribution during 2 h of continuous 1-Hz stimulation (Fig. 6 B, right).


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)

The subcellular distribution of CaM-YFP was not changed by 10- or 1-Hz electrical stimulation. (A) A typical living fiber expressing CaM-YFP is shown for 30 min (−30 and 0) before stimulation and 60 and 120 min after repetitive stimulation with 10-Hz trains. In resting fibers, CaM-YFP was present in the cytoplasm as well as the nucleus. (B) The fluorescence signals from both the nucleus and the cytoplasm were quantitated. Data are presented as the ratio of the average fluorescence intensity per pixel from the nucleus relative to the cytoplasm. Neither 10-Hz train stimulation (left) for 2 h nor 1-Hz continuous stimulation (right) resulted in any subcellular redistribution of CaM-YFP. (C) FRAP of CaM-YFP was performed in the nuclear area of a muscle fiber. Shown is a nucleus before photobleaching, immediately after photobleaching, and after 30 and 60 min of recovery. The entire nuclear region was selectively bleached. Bars, 10 μm.
© Copyright Policy
Related In: Results  -  Collection

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

fig6: The subcellular distribution of CaM-YFP was not changed by 10- or 1-Hz electrical stimulation. (A) A typical living fiber expressing CaM-YFP is shown for 30 min (−30 and 0) before stimulation and 60 and 120 min after repetitive stimulation with 10-Hz trains. In resting fibers, CaM-YFP was present in the cytoplasm as well as the nucleus. (B) The fluorescence signals from both the nucleus and the cytoplasm were quantitated. Data are presented as the ratio of the average fluorescence intensity per pixel from the nucleus relative to the cytoplasm. Neither 10-Hz train stimulation (left) for 2 h nor 1-Hz continuous stimulation (right) resulted in any subcellular redistribution of CaM-YFP. (C) FRAP of CaM-YFP was performed in the nuclear area of a muscle fiber. Shown is a nucleus before photobleaching, immediately after photobleaching, and after 30 and 60 min of recovery. The entire nuclear region was selectively bleached. Bars, 10 μm.
Mentions: Because CaMK appears to mediate the stimulation-dependent translocation of HDAC4 from the nucleus to the cytoplasm (Fig. 2), we next examined the intracellular distribution of CaM-YFP, expressed by adenoviral infection, to see if stimulation recruited CaM to the nucleus. In resting fibers, CaM-YFP exhibited a sarcomeric pattern in the cytoplasm, with the bright lines in the pattern corresponding to α-actinin localization at the Z line (not depicted), and a diffuse pattern in the nucleus except the nucleolus (Fig. 6 A). There was significant CaM-YFP in the A-band region between Z-lines, giving a pixel fluorescence about half that at the Z-line and indicating presence of CaM within the sarcomere away from the Z-line. The mean value of the ratio of nuclear to cytoplasmic mean pixel fluorescence was 0.79 ± 0.05 (11 nuclei from 7 fibers). This ratio was stable in resting fibers and did not change during a 2-h period of repetitive stimulation with 5-s trains of 10-Hz stimuli applied every 50 s (Fig. 6 B, left), a stimulus pattern that produced clear KN-62-sensitive, stimulation-dependent translocation of HDAC4 from the nucleus to the cytoplasm (Fig. 2). CaM-YFP also did not exhibit any change in nuclear-cytoplasmic distribution during 2 h of continuous 1-Hz stimulation (Fig. 6 B, right).

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