Orientation of myosin binding protein C in the cardiac muscle sarcomere determined by domain-specific immuno-EM

Kyounghwan Lee, Samantha P. Harris, Sakthivel Sadayappan, Roger Craig

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Myosin binding protein C is a thick filament protein of vertebrate striated muscle. The cardiac isoform [cardiac myosin binding protein C (cMyBP-C)] is essential for normal cardiac function, and mutations in cMyBP-C cause cardiac muscle disease. The rod-shaped molecule is composed primarily of 11 immunoglobulin- or fibronectin-like domains and is located at nine sites, 43 nm apart, in each half of the A-band. To understand how cMyBP-C functions, it is important to know its structural organization in the sarcomere, as this will affect its ability to interact with other sarcomeric proteins. Several models, in which cMyBP-C wraps around, extends radially from, or runs axially along the thick filament, have been proposed. Our goal was to define cMyBP-C orientation by determining the relative axial positions of different cMyBP-C domains. Immuno-electron microscopy was performed using mouse cardiac myofibrils labeled with antibodies specific to the N- and C-terminal domains and to the middle of cMyBP-C. Antibodies to all regions of the molecule, except the C-terminus, labeled at the same nine axial positions in each half A-band, consistent with a circumferential and/or radial rather than an axial orientation of the bulk of the molecule. The C-terminal antibody stripes were slightly displaced axially, demonstrating an axial orientation of the C-terminal three domains, with the C-terminus closer to the M-line. These results, combined with previous studies, suggest that the C-terminal domains of cMyBP-C run along the thick filament surface, while the N-terminus extends toward neighboring thin filaments. This organization provides a structural framework for understanding cMyBP-C's modulation of cardiac muscle contraction.

Original languageEnglish (US)
Pages (from-to)274-286
Number of pages13
JournalJournal of Molecular Biology
Volume427
Issue number2
DOIs
StatePublished - Jan 30 2015

Fingerprint

Sarcomeres
Cardiac Myosins
Myocardium
Antibodies
Striated Muscle
Immunoelectron Microscopy
myosin-binding protein C
Myofibrils
Muscle Contraction
Fibronectins
Vertebrates
Immunoglobulins
Heart Diseases
Protein Isoforms
Proteins
Mutation

Keywords

  • Cardiac muscle contraction
  • Cardiac muscle disease
  • Cardiac muscle regulation
  • Cardiac muscle structure
  • cMyBP-C

ASJC Scopus subject areas

  • Molecular Biology

Cite this

Orientation of myosin binding protein C in the cardiac muscle sarcomere determined by domain-specific immuno-EM. / Lee, Kyounghwan; Harris, Samantha P.; Sadayappan, Sakthivel; Craig, Roger.

In: Journal of Molecular Biology, Vol. 427, No. 2, 30.01.2015, p. 274-286.

Research output: Contribution to journalArticle

Lee, Kyounghwan ; Harris, Samantha P. ; Sadayappan, Sakthivel ; Craig, Roger. / Orientation of myosin binding protein C in the cardiac muscle sarcomere determined by domain-specific immuno-EM. In: Journal of Molecular Biology. 2015 ; Vol. 427, No. 2. pp. 274-286.
@article{e7bf6239d6b54d17b9328e112342e1fc,
title = "Orientation of myosin binding protein C in the cardiac muscle sarcomere determined by domain-specific immuno-EM",
abstract = "Myosin binding protein C is a thick filament protein of vertebrate striated muscle. The cardiac isoform [cardiac myosin binding protein C (cMyBP-C)] is essential for normal cardiac function, and mutations in cMyBP-C cause cardiac muscle disease. The rod-shaped molecule is composed primarily of 11 immunoglobulin- or fibronectin-like domains and is located at nine sites, 43 nm apart, in each half of the A-band. To understand how cMyBP-C functions, it is important to know its structural organization in the sarcomere, as this will affect its ability to interact with other sarcomeric proteins. Several models, in which cMyBP-C wraps around, extends radially from, or runs axially along the thick filament, have been proposed. Our goal was to define cMyBP-C orientation by determining the relative axial positions of different cMyBP-C domains. Immuno-electron microscopy was performed using mouse cardiac myofibrils labeled with antibodies specific to the N- and C-terminal domains and to the middle of cMyBP-C. Antibodies to all regions of the molecule, except the C-terminus, labeled at the same nine axial positions in each half A-band, consistent with a circumferential and/or radial rather than an axial orientation of the bulk of the molecule. The C-terminal antibody stripes were slightly displaced axially, demonstrating an axial orientation of the C-terminal three domains, with the C-terminus closer to the M-line. These results, combined with previous studies, suggest that the C-terminal domains of cMyBP-C run along the thick filament surface, while the N-terminus extends toward neighboring thin filaments. This organization provides a structural framework for understanding cMyBP-C's modulation of cardiac muscle contraction.",
keywords = "Cardiac muscle contraction, Cardiac muscle disease, Cardiac muscle regulation, Cardiac muscle structure, cMyBP-C",
author = "Kyounghwan Lee and Harris, {Samantha P.} and Sakthivel Sadayappan and Roger Craig",
year = "2015",
month = "1",
day = "30",
doi = "10.1016/j.jmb.2014.10.023",
language = "English (US)",
volume = "427",
pages = "274--286",
journal = "Journal of Molecular Biology",
issn = "0022-2836",
publisher = "Academic Press Inc.",
number = "2",

}

TY - JOUR

T1 - Orientation of myosin binding protein C in the cardiac muscle sarcomere determined by domain-specific immuno-EM

AU - Lee, Kyounghwan

AU - Harris, Samantha P.

AU - Sadayappan, Sakthivel

AU - Craig, Roger

PY - 2015/1/30

Y1 - 2015/1/30

N2 - Myosin binding protein C is a thick filament protein of vertebrate striated muscle. The cardiac isoform [cardiac myosin binding protein C (cMyBP-C)] is essential for normal cardiac function, and mutations in cMyBP-C cause cardiac muscle disease. The rod-shaped molecule is composed primarily of 11 immunoglobulin- or fibronectin-like domains and is located at nine sites, 43 nm apart, in each half of the A-band. To understand how cMyBP-C functions, it is important to know its structural organization in the sarcomere, as this will affect its ability to interact with other sarcomeric proteins. Several models, in which cMyBP-C wraps around, extends radially from, or runs axially along the thick filament, have been proposed. Our goal was to define cMyBP-C orientation by determining the relative axial positions of different cMyBP-C domains. Immuno-electron microscopy was performed using mouse cardiac myofibrils labeled with antibodies specific to the N- and C-terminal domains and to the middle of cMyBP-C. Antibodies to all regions of the molecule, except the C-terminus, labeled at the same nine axial positions in each half A-band, consistent with a circumferential and/or radial rather than an axial orientation of the bulk of the molecule. The C-terminal antibody stripes were slightly displaced axially, demonstrating an axial orientation of the C-terminal three domains, with the C-terminus closer to the M-line. These results, combined with previous studies, suggest that the C-terminal domains of cMyBP-C run along the thick filament surface, while the N-terminus extends toward neighboring thin filaments. This organization provides a structural framework for understanding cMyBP-C's modulation of cardiac muscle contraction.

AB - Myosin binding protein C is a thick filament protein of vertebrate striated muscle. The cardiac isoform [cardiac myosin binding protein C (cMyBP-C)] is essential for normal cardiac function, and mutations in cMyBP-C cause cardiac muscle disease. The rod-shaped molecule is composed primarily of 11 immunoglobulin- or fibronectin-like domains and is located at nine sites, 43 nm apart, in each half of the A-band. To understand how cMyBP-C functions, it is important to know its structural organization in the sarcomere, as this will affect its ability to interact with other sarcomeric proteins. Several models, in which cMyBP-C wraps around, extends radially from, or runs axially along the thick filament, have been proposed. Our goal was to define cMyBP-C orientation by determining the relative axial positions of different cMyBP-C domains. Immuno-electron microscopy was performed using mouse cardiac myofibrils labeled with antibodies specific to the N- and C-terminal domains and to the middle of cMyBP-C. Antibodies to all regions of the molecule, except the C-terminus, labeled at the same nine axial positions in each half A-band, consistent with a circumferential and/or radial rather than an axial orientation of the bulk of the molecule. The C-terminal antibody stripes were slightly displaced axially, demonstrating an axial orientation of the C-terminal three domains, with the C-terminus closer to the M-line. These results, combined with previous studies, suggest that the C-terminal domains of cMyBP-C run along the thick filament surface, while the N-terminus extends toward neighboring thin filaments. This organization provides a structural framework for understanding cMyBP-C's modulation of cardiac muscle contraction.

KW - Cardiac muscle contraction

KW - Cardiac muscle disease

KW - Cardiac muscle regulation

KW - Cardiac muscle structure

KW - cMyBP-C

UR - http://www.scopus.com/inward/record.url?scp=84920784546&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84920784546&partnerID=8YFLogxK

U2 - 10.1016/j.jmb.2014.10.023

DO - 10.1016/j.jmb.2014.10.023

M3 - Article

VL - 427

SP - 274

EP - 286

JO - Journal of Molecular Biology

JF - Journal of Molecular Biology

SN - 0022-2836

IS - 2

ER -