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Contribution of the myosin binding protein C motif to functional effects in permeabilized rat trabeculae.

Razumova MV, Bezold KL, Tu AY, Regnier M, Harris SP - J. Gen. Physiol. (2008)

Bottom Line: Myosin binding protein C (MyBP-C) is a thick-filament protein that limits cross-bridge cycling rates and reduces myocyte power output.Recombinant proteins that lacked the combination of C1 and the motif did not affect contractile properties.These results suggest that the C1 domain plus the motif constitute a functional unit of MyBP-C that can activate the thin filament.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, University of Washington, Seattle, WA 98195, USA.

ABSTRACT
Myosin binding protein C (MyBP-C) is a thick-filament protein that limits cross-bridge cycling rates and reduces myocyte power output. To investigate mechanisms by which MyBP-C affects contraction, we assessed effects of recombinant N-terminal domains of cardiac MyBP-C (cMyBP-C) on contractile properties of permeabilized rat cardiac trabeculae. Here, we show that N-terminal fragments of cMyBP-C that contained the first three immunoglobulin domains of cMyBP-C (i.e., C0, C1, and C2) plus the unique linker sequence termed the MyBP-C "motif" or "m-domain" increased Ca(2+) sensitivity of tension and increased rates of tension redevelopment (i.e., k(tr)) at submaximal levels of Ca(2+). At concentrations > or =20 microM, recombinant proteins also activated force in the absence of Ca(2+) and inhibited maximum Ca(2+)-activated force. Recombinant proteins that lacked the combination of C1 and the motif did not affect contractile properties. These results suggest that the C1 domain plus the motif constitute a functional unit of MyBP-C that can activate the thin filament.

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Effects of the MyBP-C motif on Ca2+ sensitivity of tension. (A) Incubation of trabeculae with 10 μM C1C2 (squares; n = 6) resulted in a leftward shift of the tension–pCa relationship relative to control (circles), indicating an increase in Ca2+ sensitivity of tension (ΔpCa50 = 0.30 ± 0.05). 20 μM C1C2(-m) (triangles; n = 3) that lacks the MyBP-C motif was without effect on the tension–pCa relationship. (B) 20 μM C2C4 without the motif (triangles; n = 4) and 20 μM C3mC4 with the motif (squares; n = 3) had no effect on Ca2+ sensitivity of tension.
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fig3: Effects of the MyBP-C motif on Ca2+ sensitivity of tension. (A) Incubation of trabeculae with 10 μM C1C2 (squares; n = 6) resulted in a leftward shift of the tension–pCa relationship relative to control (circles), indicating an increase in Ca2+ sensitivity of tension (ΔpCa50 = 0.30 ± 0.05). 20 μM C1C2(-m) (triangles; n = 3) that lacks the MyBP-C motif was without effect on the tension–pCa relationship. (B) 20 μM C2C4 without the motif (triangles; n = 4) and 20 μM C3mC4 with the motif (squares; n = 3) had no effect on Ca2+ sensitivity of tension.

Mentions: We had previously shown that a recombinant protein containing the C1 through C2 domains of cMyBP-C (i.e., C1C2; Fig. 1) increased Ca2+ sensitivity of tension in permeabilized mouse myocytes (Harris et al., 2004). To determine whether the MyBP-C motif was required for the effects of C1C2, we created a mutant C1C2 protein that lacked the motif sequence between the C1 and C2 domains (i.e., C1C2[-m]) and compared its effects to those of C1C2 (replete with motif) on the Ca2+ sensitivity of force in permeabilized rat cardiac trabeculae. As shown in Fig. 3 A, 10 μM C1C2 added to relaxing (pCa 9.0) and pre-activating solutions resulted in significantly greater Ca2+-activated force developed in submaximal pCa solutions, as indicated by the leftward shift of the force–pCa relationship compared with control curves generated before the addition of protein. These data thus agree with previous observations that C1C2 increased Ca2+ sensitivity of tension in permeabilized mouse myocytes (Harris et al., 2004). However, in marked contrast to the effects of C1C2, 20 μM C1C2(-m), which lacked the motif, did not increase Ca2+ sensitivity of tension, suggesting that the motif is required for the effects of C1C2 on force at submaximal pCa concentrations.


Contribution of the myosin binding protein C motif to functional effects in permeabilized rat trabeculae.

Razumova MV, Bezold KL, Tu AY, Regnier M, Harris SP - J. Gen. Physiol. (2008)

Effects of the MyBP-C motif on Ca2+ sensitivity of tension. (A) Incubation of trabeculae with 10 μM C1C2 (squares; n = 6) resulted in a leftward shift of the tension–pCa relationship relative to control (circles), indicating an increase in Ca2+ sensitivity of tension (ΔpCa50 = 0.30 ± 0.05). 20 μM C1C2(-m) (triangles; n = 3) that lacks the MyBP-C motif was without effect on the tension–pCa relationship. (B) 20 μM C2C4 without the motif (triangles; n = 4) and 20 μM C3mC4 with the motif (squares; n = 3) had no effect on Ca2+ sensitivity of tension.
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2571974&req=5

fig3: Effects of the MyBP-C motif on Ca2+ sensitivity of tension. (A) Incubation of trabeculae with 10 μM C1C2 (squares; n = 6) resulted in a leftward shift of the tension–pCa relationship relative to control (circles), indicating an increase in Ca2+ sensitivity of tension (ΔpCa50 = 0.30 ± 0.05). 20 μM C1C2(-m) (triangles; n = 3) that lacks the MyBP-C motif was without effect on the tension–pCa relationship. (B) 20 μM C2C4 without the motif (triangles; n = 4) and 20 μM C3mC4 with the motif (squares; n = 3) had no effect on Ca2+ sensitivity of tension.
Mentions: We had previously shown that a recombinant protein containing the C1 through C2 domains of cMyBP-C (i.e., C1C2; Fig. 1) increased Ca2+ sensitivity of tension in permeabilized mouse myocytes (Harris et al., 2004). To determine whether the MyBP-C motif was required for the effects of C1C2, we created a mutant C1C2 protein that lacked the motif sequence between the C1 and C2 domains (i.e., C1C2[-m]) and compared its effects to those of C1C2 (replete with motif) on the Ca2+ sensitivity of force in permeabilized rat cardiac trabeculae. As shown in Fig. 3 A, 10 μM C1C2 added to relaxing (pCa 9.0) and pre-activating solutions resulted in significantly greater Ca2+-activated force developed in submaximal pCa solutions, as indicated by the leftward shift of the force–pCa relationship compared with control curves generated before the addition of protein. These data thus agree with previous observations that C1C2 increased Ca2+ sensitivity of tension in permeabilized mouse myocytes (Harris et al., 2004). However, in marked contrast to the effects of C1C2, 20 μM C1C2(-m), which lacked the motif, did not increase Ca2+ sensitivity of tension, suggesting that the motif is required for the effects of C1C2 on force at submaximal pCa concentrations.

Bottom Line: Myosin binding protein C (MyBP-C) is a thick-filament protein that limits cross-bridge cycling rates and reduces myocyte power output.Recombinant proteins that lacked the combination of C1 and the motif did not affect contractile properties.These results suggest that the C1 domain plus the motif constitute a functional unit of MyBP-C that can activate the thin filament.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, University of Washington, Seattle, WA 98195, USA.

ABSTRACT
Myosin binding protein C (MyBP-C) is a thick-filament protein that limits cross-bridge cycling rates and reduces myocyte power output. To investigate mechanisms by which MyBP-C affects contraction, we assessed effects of recombinant N-terminal domains of cardiac MyBP-C (cMyBP-C) on contractile properties of permeabilized rat cardiac trabeculae. Here, we show that N-terminal fragments of cMyBP-C that contained the first three immunoglobulin domains of cMyBP-C (i.e., C0, C1, and C2) plus the unique linker sequence termed the MyBP-C "motif" or "m-domain" increased Ca(2+) sensitivity of tension and increased rates of tension redevelopment (i.e., k(tr)) at submaximal levels of Ca(2+). At concentrations > or =20 microM, recombinant proteins also activated force in the absence of Ca(2+) and inhibited maximum Ca(2+)-activated force. Recombinant proteins that lacked the combination of C1 and the motif did not affect contractile properties. These results suggest that the C1 domain plus the motif constitute a functional unit of MyBP-C that can activate the thin filament.

Show MeSH
Related in: MedlinePlus