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Localization and regulation of the N terminal splice variant of PGC-1α in adult skeletal muscle fibers.

Shen T, Liu Y, Schneider MF - J. Biomed. Biotechnol. (2012)

Bottom Line: Recently, a novel splice variant of PGC-1α (NT-PGC-1α, amino acids 1-270) was cloned and found to be expressed in muscle.Activation of p38 MAPK by muscle activity or of AMPK had no effect on the subcellular distribution of NT-PGC-1α.Together these results suggest that the regulation of NT-PGC-1α in muscle fibers may be very different from that of the full-length PGC-1α, which is exclusively nuclear.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, MD 21201-1503, USA.

ABSTRACT
The transcriptional coactivator peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) regulates expression of genes for metabolism and muscle fiber type. Recently, a novel splice variant of PGC-1α (NT-PGC-1α, amino acids 1-270) was cloned and found to be expressed in muscle. Here we use Flag-tagged NT-PGC-1α to examine the subcellular localization and regulation of NT-PGC-1α in skeletal muscle fibers. Flag-NT-PGC-1α is located predominantly in the myoplasm. Nuclear NT-PGC-1α can be increased by activation of protein kinase A. Activation of p38 MAPK by muscle activity or of AMPK had no effect on the subcellular distribution of NT-PGC-1α. Inhibition of CRM1-mediated export only caused relatively slow nuclear accumulation of NT-PGC-1α, indicating that nuclear export of NT-PGC-1α may be mediated by both CRM1-dependent and -independent pathways. Together these results suggest that the regulation of NT-PGC-1α in muscle fibers may be very different from that of the full-length PGC-1α, which is exclusively nuclear.

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Activation of PKA increases nuclear Flag-NT-PGC-1α. (a) Representative images of Flag-NT-PGC-1α in a control fiber (control) and in another fiber after 1 h treatment with 1 mM dbcAMP in culture medium in the tissue culture incubator (dbcAMP). Scale bar, 10 μm. (b) The n/c fluorescence ratio of Flag-NT-PGC-1α in muscle fibers with (dbcAMP) or without (control) dbcAMP treatment. n/c values from 20 nuclei from 20 randomly selected fibers were averaged to give the mean value for each group. Asterisk indicates statistical significance between groups at P < 0.05.
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fig2: Activation of PKA increases nuclear Flag-NT-PGC-1α. (a) Representative images of Flag-NT-PGC-1α in a control fiber (control) and in another fiber after 1 h treatment with 1 mM dbcAMP in culture medium in the tissue culture incubator (dbcAMP). Scale bar, 10 μm. (b) The n/c fluorescence ratio of Flag-NT-PGC-1α in muscle fibers with (dbcAMP) or without (control) dbcAMP treatment. n/c values from 20 nuclei from 20 randomly selected fibers were averaged to give the mean value for each group. Asterisk indicates statistical significance between groups at P < 0.05.

Mentions: Recent studies in adipocytes and CHO-K1 cells have shown that activation of PKA can significantly increase the nuclear content of NT-PGC-1α [12, 13]. To test whether this regulation mechanism also exists in skeletal muscle fibers, we treated muscle fibers with 1 mM dbcAMP for 1 h in culture medium in the tissue culture incubator, as previously used to activate PKA in CHO-K1 cells [13]. Our results show that the Flag-NT-PGC-1α fluorescent staining was stronger in nuclei of fibers exposed to dbcAMP (Figure 2(a), bdcAMP) than in control fibers (Figure 2(a), control). The Flag-NT-PGC-1α n/c fluorescence ratio was significantly increased from 0.71 ± 0.03 in control to 0.98 ± 0.07 in dbcAMP-treated fibers (Figure 2(b); P < 0.05), a 1.4-fold increase. The nuclear increase of Flag-NT-PGC-1α after 1 h treatment of dbcAMP found here in skeletal muscle fibers is very close to what has been previously obtained in CHO-K1 cells (about 1.4 fold) and in differentiated adipocytes (about 1.5 fold) [13]. Together, these results support the notion that PKA regulates the subcellular distribution of NT-PGC-1α and is thereby involved in modulation of the NT-PGC-1α-dependent signaling pathway in muscle fibers. In brown adipocytes, it has been shown that PKA-dependent regulation of nuclear content of NT-PGC-1α plays a role in the transcriptional activation of UCP1 and CIDEA [13], but the downstream signaling pathway of NT-PGC-1α in skeletal muscle remains to be determined.


Localization and regulation of the N terminal splice variant of PGC-1α in adult skeletal muscle fibers.

Shen T, Liu Y, Schneider MF - J. Biomed. Biotechnol. (2012)

Activation of PKA increases nuclear Flag-NT-PGC-1α. (a) Representative images of Flag-NT-PGC-1α in a control fiber (control) and in another fiber after 1 h treatment with 1 mM dbcAMP in culture medium in the tissue culture incubator (dbcAMP). Scale bar, 10 μm. (b) The n/c fluorescence ratio of Flag-NT-PGC-1α in muscle fibers with (dbcAMP) or without (control) dbcAMP treatment. n/c values from 20 nuclei from 20 randomly selected fibers were averaged to give the mean value for each group. Asterisk indicates statistical significance between groups at P < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3303705&req=5

fig2: Activation of PKA increases nuclear Flag-NT-PGC-1α. (a) Representative images of Flag-NT-PGC-1α in a control fiber (control) and in another fiber after 1 h treatment with 1 mM dbcAMP in culture medium in the tissue culture incubator (dbcAMP). Scale bar, 10 μm. (b) The n/c fluorescence ratio of Flag-NT-PGC-1α in muscle fibers with (dbcAMP) or without (control) dbcAMP treatment. n/c values from 20 nuclei from 20 randomly selected fibers were averaged to give the mean value for each group. Asterisk indicates statistical significance between groups at P < 0.05.
Mentions: Recent studies in adipocytes and CHO-K1 cells have shown that activation of PKA can significantly increase the nuclear content of NT-PGC-1α [12, 13]. To test whether this regulation mechanism also exists in skeletal muscle fibers, we treated muscle fibers with 1 mM dbcAMP for 1 h in culture medium in the tissue culture incubator, as previously used to activate PKA in CHO-K1 cells [13]. Our results show that the Flag-NT-PGC-1α fluorescent staining was stronger in nuclei of fibers exposed to dbcAMP (Figure 2(a), bdcAMP) than in control fibers (Figure 2(a), control). The Flag-NT-PGC-1α n/c fluorescence ratio was significantly increased from 0.71 ± 0.03 in control to 0.98 ± 0.07 in dbcAMP-treated fibers (Figure 2(b); P < 0.05), a 1.4-fold increase. The nuclear increase of Flag-NT-PGC-1α after 1 h treatment of dbcAMP found here in skeletal muscle fibers is very close to what has been previously obtained in CHO-K1 cells (about 1.4 fold) and in differentiated adipocytes (about 1.5 fold) [13]. Together, these results support the notion that PKA regulates the subcellular distribution of NT-PGC-1α and is thereby involved in modulation of the NT-PGC-1α-dependent signaling pathway in muscle fibers. In brown adipocytes, it has been shown that PKA-dependent regulation of nuclear content of NT-PGC-1α plays a role in the transcriptional activation of UCP1 and CIDEA [13], but the downstream signaling pathway of NT-PGC-1α in skeletal muscle remains to be determined.

Bottom Line: Recently, a novel splice variant of PGC-1α (NT-PGC-1α, amino acids 1-270) was cloned and found to be expressed in muscle.Activation of p38 MAPK by muscle activity or of AMPK had no effect on the subcellular distribution of NT-PGC-1α.Together these results suggest that the regulation of NT-PGC-1α in muscle fibers may be very different from that of the full-length PGC-1α, which is exclusively nuclear.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, MD 21201-1503, USA.

ABSTRACT
The transcriptional coactivator peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) regulates expression of genes for metabolism and muscle fiber type. Recently, a novel splice variant of PGC-1α (NT-PGC-1α, amino acids 1-270) was cloned and found to be expressed in muscle. Here we use Flag-tagged NT-PGC-1α to examine the subcellular localization and regulation of NT-PGC-1α in skeletal muscle fibers. Flag-NT-PGC-1α is located predominantly in the myoplasm. Nuclear NT-PGC-1α can be increased by activation of protein kinase A. Activation of p38 MAPK by muscle activity or of AMPK had no effect on the subcellular distribution of NT-PGC-1α. Inhibition of CRM1-mediated export only caused relatively slow nuclear accumulation of NT-PGC-1α, indicating that nuclear export of NT-PGC-1α may be mediated by both CRM1-dependent and -independent pathways. Together these results suggest that the regulation of NT-PGC-1α in muscle fibers may be very different from that of the full-length PGC-1α, which is exclusively nuclear.

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