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New Isoform of Cardiac Myosin Light Chain Kinase and the Role of Cardiac Myosin Phosphorylation in α1-Adrenoceptor Mediated Inotropic Response.

Taniguchi M, Okamoto R, Ito M, Goto I, Fujita S, Konishi K, Mizutani H, Dohi K, Hartshorne DJ, Itoh T - PLoS ONE (2015)

Bottom Line: The levels of MLC2v phosphorylation was significantly lower in C57BL/6N than in C57BL/6J.Phenylephrine had no effect on MLC2v phosphorylation in either strains but increased the twitch tension more potently in C57BL/6J than in C57BL/6N.There was a significant positive correlation between the phenylephrine-induced inotropic response and the levels of MLC2v phosphorylation within ranges of 15-30%.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology and Nephrology, Mie University Graduate School of Medicine, Edobashi, Tsu 514-8507, Mie, Japan.

ABSTRACT

Background & aims: Cardiac myosin light chain kinase (cMLCK) plays an obligatory role in maintaining the phosphorylation levels of regulatory myosin light chain (MLC2), which is thought to be crucial for regulation of cardiac function. To test this hypothesis, the role played by ventricular MLC2 (MLC2v) phosphorylation was investigated in the phenylephrine-induced increase in twitch tension using the naturally-occurring mouse strain, C57BL/6N, in which cMLCK is down regulated.

Methods and results: By Western blot and nanoLC-MS/MS analysis, cMLCKs with molecular mass of 61-kDa (cMLCK-2) and/or 86-kDa were identified in mice heart. Among various mouse strains, C57BL/6N expressed cMLCK-2 alone and the closest relative strain C57BL/6J expressed both cMLCKs. The levels of MLC2v phosphorylation was significantly lower in C57BL/6N than in C57BL/6J. The papillary muscle twitch tension induced by electrical field stimulation was smaller in C57BL/6N than C57BL/6J. Phenylephrine had no effect on MLC2v phosphorylation in either strains but increased the twitch tension more potently in C57BL/6J than in C57BL/6N. Calyculin A increased papillary muscle MLC2v phosphorylation to a similar extent in both strains but increased the phenylephrine-induced inotropic response only in C57BL/6N. There was a significant positive correlation between the phenylephrine-induced inotropic response and the levels of MLC2v phosphorylation within ranges of 15-30%.

Conclusions: We identified a new isoform of cMLCK with a molecular mass of 61kDa(cMLCK-2) in mouse heart. In the C57BL/6N strain, only cMLCK-2 was expressed and the basal MLC2v phosphorylation levels and the phenylephrine-induced inotropic response were both smaller. We suggest that a lower phenylephrine-induced inotropic response may be caused by the lower basal MLC2v phosphorylation levels in this strain.

No MeSH data available.


Related in: MedlinePlus

Effects of phenylephrine (PE) on twitch tension and MLC2v phosphorylation in papillary muscles from C57BL/6J and C57BL/6N.(A) Representative tension recordings. (B) The phosphorylation levels of MLC2v. The % ratio of phosphorylated MLC2v vs. total MLC2v was calculated (n = 5). (C) Concentration-dependent effects of phenylephrine in twitch tension. After basal twitch tension was recorded, phenylephrine was cumulatively applied. Atenolol was present throughout the experiments for tension measurements. *P<0.05, **P<0.01, ***P<0.001, n = 5–6.
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pone.0141130.g005: Effects of phenylephrine (PE) on twitch tension and MLC2v phosphorylation in papillary muscles from C57BL/6J and C57BL/6N.(A) Representative tension recordings. (B) The phosphorylation levels of MLC2v. The % ratio of phosphorylated MLC2v vs. total MLC2v was calculated (n = 5). (C) Concentration-dependent effects of phenylephrine in twitch tension. After basal twitch tension was recorded, phenylephrine was cumulatively applied. Atenolol was present throughout the experiments for tension measurements. *P<0.05, **P<0.01, ***P<0.001, n = 5–6.

Mentions: Phenylephrine did not modify MLC2v phosphorylation in either C57BL/6J or C57BL/6N but it enhanced the amplitude of twitch tension in both strains (Fig 5A and 5B). Inotropic action of phenylephrine was more pronounced in C57BL/6J than in C57BL/6N (Fig 5C). No significant changes in MLC2v phosphorylation by phenylephrine were confirmed by using anti-phospho-MLC antibody (data not shown).


New Isoform of Cardiac Myosin Light Chain Kinase and the Role of Cardiac Myosin Phosphorylation in α1-Adrenoceptor Mediated Inotropic Response.

Taniguchi M, Okamoto R, Ito M, Goto I, Fujita S, Konishi K, Mizutani H, Dohi K, Hartshorne DJ, Itoh T - PLoS ONE (2015)

Effects of phenylephrine (PE) on twitch tension and MLC2v phosphorylation in papillary muscles from C57BL/6J and C57BL/6N.(A) Representative tension recordings. (B) The phosphorylation levels of MLC2v. The % ratio of phosphorylated MLC2v vs. total MLC2v was calculated (n = 5). (C) Concentration-dependent effects of phenylephrine in twitch tension. After basal twitch tension was recorded, phenylephrine was cumulatively applied. Atenolol was present throughout the experiments for tension measurements. *P<0.05, **P<0.01, ***P<0.001, n = 5–6.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4626101&req=5

pone.0141130.g005: Effects of phenylephrine (PE) on twitch tension and MLC2v phosphorylation in papillary muscles from C57BL/6J and C57BL/6N.(A) Representative tension recordings. (B) The phosphorylation levels of MLC2v. The % ratio of phosphorylated MLC2v vs. total MLC2v was calculated (n = 5). (C) Concentration-dependent effects of phenylephrine in twitch tension. After basal twitch tension was recorded, phenylephrine was cumulatively applied. Atenolol was present throughout the experiments for tension measurements. *P<0.05, **P<0.01, ***P<0.001, n = 5–6.
Mentions: Phenylephrine did not modify MLC2v phosphorylation in either C57BL/6J or C57BL/6N but it enhanced the amplitude of twitch tension in both strains (Fig 5A and 5B). Inotropic action of phenylephrine was more pronounced in C57BL/6J than in C57BL/6N (Fig 5C). No significant changes in MLC2v phosphorylation by phenylephrine were confirmed by using anti-phospho-MLC antibody (data not shown).

Bottom Line: The levels of MLC2v phosphorylation was significantly lower in C57BL/6N than in C57BL/6J.Phenylephrine had no effect on MLC2v phosphorylation in either strains but increased the twitch tension more potently in C57BL/6J than in C57BL/6N.There was a significant positive correlation between the phenylephrine-induced inotropic response and the levels of MLC2v phosphorylation within ranges of 15-30%.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology and Nephrology, Mie University Graduate School of Medicine, Edobashi, Tsu 514-8507, Mie, Japan.

ABSTRACT

Background & aims: Cardiac myosin light chain kinase (cMLCK) plays an obligatory role in maintaining the phosphorylation levels of regulatory myosin light chain (MLC2), which is thought to be crucial for regulation of cardiac function. To test this hypothesis, the role played by ventricular MLC2 (MLC2v) phosphorylation was investigated in the phenylephrine-induced increase in twitch tension using the naturally-occurring mouse strain, C57BL/6N, in which cMLCK is down regulated.

Methods and results: By Western blot and nanoLC-MS/MS analysis, cMLCKs with molecular mass of 61-kDa (cMLCK-2) and/or 86-kDa were identified in mice heart. Among various mouse strains, C57BL/6N expressed cMLCK-2 alone and the closest relative strain C57BL/6J expressed both cMLCKs. The levels of MLC2v phosphorylation was significantly lower in C57BL/6N than in C57BL/6J. The papillary muscle twitch tension induced by electrical field stimulation was smaller in C57BL/6N than C57BL/6J. Phenylephrine had no effect on MLC2v phosphorylation in either strains but increased the twitch tension more potently in C57BL/6J than in C57BL/6N. Calyculin A increased papillary muscle MLC2v phosphorylation to a similar extent in both strains but increased the phenylephrine-induced inotropic response only in C57BL/6N. There was a significant positive correlation between the phenylephrine-induced inotropic response and the levels of MLC2v phosphorylation within ranges of 15-30%.

Conclusions: We identified a new isoform of cMLCK with a molecular mass of 61kDa(cMLCK-2) in mouse heart. In the C57BL/6N strain, only cMLCK-2 was expressed and the basal MLC2v phosphorylation levels and the phenylephrine-induced inotropic response were both smaller. We suggest that a lower phenylephrine-induced inotropic response may be caused by the lower basal MLC2v phosphorylation levels in this strain.

No MeSH data available.


Related in: MedlinePlus