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Repression of slow myosin heavy chain 2 gene expression in fast skeletal muscle fibers by muscarinic acetylcholine receptor and G(alpha)q signaling.

Jordan T, Li J, Jiang H, DiMario JX - J. Cell Biol. (2003)

Bottom Line: Increased G(alpha)q activity repressed slow MyHC2 expression to nondetectable levels in innervated MA fibers.Decreased PKC activity in atropine-treated innervated PM fibers correlated with slow MyHC2 expression.These data suggest that slow MyHC2 repression in innervated fast PM fibers is mediated by cell signaling involving mAchRs, G(alpha)q, and PKC.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Anatomy, Chicago Medical School, North Chicago, IL 60064, USA.

ABSTRACT
Gene expression in skeletal muscle fibers is regulated by innervation and intrinsic fiber properties. To determine the mechanism of repression of slow MyHC2 expression in innervated fast pectoralis major (PM) fibers, we investigated the function of the muscarinic acetylcholine receptor (mAchR) and G(alpha)q. Both mAchR and G(alpha)q are abundant in medial adductor (MA) and PM fibers, and mAchR and G(alpha)q interact in these fibers. Whereas innervation of PM fibers was insufficient to induce slow MyHC2 expression, inhibition of mAchR activity with atropine in innervated PM fibers induced slow MyHC2 expression. Increased G(alpha)q activity repressed slow MyHC2 expression to nondetectable levels in innervated MA fibers. Reduced mAchR activity decreased PKC activity in PM fibers, and increased G(alpha)q activity increased PKC activity in PM and MA fibers. Decreased PKC activity in atropine-treated innervated PM fibers correlated with slow MyHC2 expression. These data suggest that slow MyHC2 repression in innervated fast PM fibers is mediated by cell signaling involving mAchRs, G(alpha)q, and PKC.

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Western blot analysis of Gαq and mAchR in PM and MA muscle. Protein extracts from ED13 PM and MA muscles were prepared, electrophoresed, and blotted as described in Materials and methods. Blots were incubated with Gαq, mAchR, and β-actin–specific antibodies and HRP-conjugated secondary antibodies. Proteins were visualized by chemiluminescence. Both PM and MA contain readily detectable amounts of Gαq and mAchR. Equal amounts (100 μg) of PM and MA extracts were loaded as determined by BCA protein assays and detection of β-actin.
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fig1: Western blot analysis of Gαq and mAchR in PM and MA muscle. Protein extracts from ED13 PM and MA muscles were prepared, electrophoresed, and blotted as described in Materials and methods. Blots were incubated with Gαq, mAchR, and β-actin–specific antibodies and HRP-conjugated secondary antibodies. Proteins were visualized by chemiluminescence. Both PM and MA contain readily detectable amounts of Gαq and mAchR. Equal amounts (100 μg) of PM and MA extracts were loaded as determined by BCA protein assays and detection of β-actin.

Mentions: Based on previous results demonstrating that expression of slow MyHC2 was regulated by PKC activity (DiMario and Funk, 1999), we hypothesized that upstream cellular signaling molecules such as the mAchR and Gαq may be present as potential regulators of PKC activity in PM and MA muscle fibers. To determine whether chicken PM and MA muscle express mAchR and Gαq, Western blots were performed on whole muscle cell extracts using M1 type mAchR and Gαq antibodies (Fig. 1). Both PM and MA contained readily detectable mAchR and Gαq in approximately equal amounts. These results are in agreement with studies of murine skeletal muscle in which Gαq and Gα11 mRNAs were present, and Gα14 and Gα15 mRNAs were absent (Strathmann and Simon, 1990; Wilkie et al., 1991).


Repression of slow myosin heavy chain 2 gene expression in fast skeletal muscle fibers by muscarinic acetylcholine receptor and G(alpha)q signaling.

Jordan T, Li J, Jiang H, DiMario JX - J. Cell Biol. (2003)

Western blot analysis of Gαq and mAchR in PM and MA muscle. Protein extracts from ED13 PM and MA muscles were prepared, electrophoresed, and blotted as described in Materials and methods. Blots were incubated with Gαq, mAchR, and β-actin–specific antibodies and HRP-conjugated secondary antibodies. Proteins were visualized by chemiluminescence. Both PM and MA contain readily detectable amounts of Gαq and mAchR. Equal amounts (100 μg) of PM and MA extracts were loaded as determined by BCA protein assays and detection of β-actin.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172810&req=5

fig1: Western blot analysis of Gαq and mAchR in PM and MA muscle. Protein extracts from ED13 PM and MA muscles were prepared, electrophoresed, and blotted as described in Materials and methods. Blots were incubated with Gαq, mAchR, and β-actin–specific antibodies and HRP-conjugated secondary antibodies. Proteins were visualized by chemiluminescence. Both PM and MA contain readily detectable amounts of Gαq and mAchR. Equal amounts (100 μg) of PM and MA extracts were loaded as determined by BCA protein assays and detection of β-actin.
Mentions: Based on previous results demonstrating that expression of slow MyHC2 was regulated by PKC activity (DiMario and Funk, 1999), we hypothesized that upstream cellular signaling molecules such as the mAchR and Gαq may be present as potential regulators of PKC activity in PM and MA muscle fibers. To determine whether chicken PM and MA muscle express mAchR and Gαq, Western blots were performed on whole muscle cell extracts using M1 type mAchR and Gαq antibodies (Fig. 1). Both PM and MA contained readily detectable mAchR and Gαq in approximately equal amounts. These results are in agreement with studies of murine skeletal muscle in which Gαq and Gα11 mRNAs were present, and Gα14 and Gα15 mRNAs were absent (Strathmann and Simon, 1990; Wilkie et al., 1991).

Bottom Line: Increased G(alpha)q activity repressed slow MyHC2 expression to nondetectable levels in innervated MA fibers.Decreased PKC activity in atropine-treated innervated PM fibers correlated with slow MyHC2 expression.These data suggest that slow MyHC2 repression in innervated fast PM fibers is mediated by cell signaling involving mAchRs, G(alpha)q, and PKC.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Anatomy, Chicago Medical School, North Chicago, IL 60064, USA.

ABSTRACT
Gene expression in skeletal muscle fibers is regulated by innervation and intrinsic fiber properties. To determine the mechanism of repression of slow MyHC2 expression in innervated fast pectoralis major (PM) fibers, we investigated the function of the muscarinic acetylcholine receptor (mAchR) and G(alpha)q. Both mAchR and G(alpha)q are abundant in medial adductor (MA) and PM fibers, and mAchR and G(alpha)q interact in these fibers. Whereas innervation of PM fibers was insufficient to induce slow MyHC2 expression, inhibition of mAchR activity with atropine in innervated PM fibers induced slow MyHC2 expression. Increased G(alpha)q activity repressed slow MyHC2 expression to nondetectable levels in innervated MA fibers. Reduced mAchR activity decreased PKC activity in PM fibers, and increased G(alpha)q activity increased PKC activity in PM and MA fibers. Decreased PKC activity in atropine-treated innervated PM fibers correlated with slow MyHC2 expression. These data suggest that slow MyHC2 repression in innervated fast PM fibers is mediated by cell signaling involving mAchRs, G(alpha)q, and PKC.

Show MeSH
Related in: MedlinePlus