Digestion of cardiac and skeletal muscle junctional sarcoplasmic reticulum vesicles with Calpain II. Effects on the Ca2+ release channel

D. P. Rardon, D. C. Cefali, R. D. Mitchell, S. M. Seiler, D. R. Hathaway, Larry Jones

Research output: Contribution to journalArticle

53 Citations (Scopus)

Abstract

The Ca2+ release channel and ryanodine receptor are activities copurifying with the 400,000-450,000 Da high molecular weight protein of cardiac and skeletal junctional sarcoplasmic reticulum. Calpain II, an endogenous cytosolic protease, was used to selectively degrade the high molecular weight protein in cardiac and skeletal muscle sarcoplasmic reticulum vesicles, and its effects on the activity of the Ca2+ release channel and [3H]ryanodine binding sites were analyzed. Degradation of the high molecular weight protein was associated with appearance of 315,000 and 150,000 Da proteolytic fragments and with a change in the ultrastructure of the 'feet', extravesicular projections that protrude from the junctional sarcoplasmic reticulum membrane. The maximal number of [3H]ryanodine binding sites and the affinities of the sites for ryanodine were not remarkably affected by calpain II. Ca2+ release channels recorded from nondegraded cardiac and skeletal membrane vesicle preparations had slope conductances of 85 and 110 pS, respectively, measured with 1 μM cis-Ca2+ and 50 mM trans-Ba2+. Proteolysis did not alter the unitary channel conductances but did increase the percentage of channel open times from 36% to more than 90%. After proteolysis, channel opening remained dependent on micromolar cis-Ca2+, and high concentrations of ryanodine (300 μM) still blocked the channel. Our results suggest that proteolysis of the Ca2+ release channel with calpain II selectively impairs its inactivation, leaving its unitary conductance and the requirement for micromolar Ca2+ intact.

Original languageEnglish
Pages (from-to)84-96
Number of pages13
JournalCirculation Research
Volume67
Issue number1
StatePublished - 1990

Fingerprint

Ryanodine
Calpain
Sarcoplasmic Reticulum
Digestion
Myocardium
Skeletal Muscle
Proteolysis
Molecular Weight
Binding Sites
Ryanodine Receptor Calcium Release Channel
Proteins
Membranes
Foot
Peptide Hydrolases

Keywords

  • calcium release channel
  • calpain
  • excitation-contraction coupling
  • sarcoplasmic reticulum

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine

Cite this

Digestion of cardiac and skeletal muscle junctional sarcoplasmic reticulum vesicles with Calpain II. Effects on the Ca2+ release channel. / Rardon, D. P.; Cefali, D. C.; Mitchell, R. D.; Seiler, S. M.; Hathaway, D. R.; Jones, Larry.

In: Circulation Research, Vol. 67, No. 1, 1990, p. 84-96.

Research output: Contribution to journalArticle

Rardon, D. P. ; Cefali, D. C. ; Mitchell, R. D. ; Seiler, S. M. ; Hathaway, D. R. ; Jones, Larry. / Digestion of cardiac and skeletal muscle junctional sarcoplasmic reticulum vesicles with Calpain II. Effects on the Ca2+ release channel. In: Circulation Research. 1990 ; Vol. 67, No. 1. pp. 84-96.
@article{bfc0289ada384909b515e10846beda8a,
title = "Digestion of cardiac and skeletal muscle junctional sarcoplasmic reticulum vesicles with Calpain II. Effects on the Ca2+ release channel",
abstract = "The Ca2+ release channel and ryanodine receptor are activities copurifying with the 400,000-450,000 Da high molecular weight protein of cardiac and skeletal junctional sarcoplasmic reticulum. Calpain II, an endogenous cytosolic protease, was used to selectively degrade the high molecular weight protein in cardiac and skeletal muscle sarcoplasmic reticulum vesicles, and its effects on the activity of the Ca2+ release channel and [3H]ryanodine binding sites were analyzed. Degradation of the high molecular weight protein was associated with appearance of 315,000 and 150,000 Da proteolytic fragments and with a change in the ultrastructure of the 'feet', extravesicular projections that protrude from the junctional sarcoplasmic reticulum membrane. The maximal number of [3H]ryanodine binding sites and the affinities of the sites for ryanodine were not remarkably affected by calpain II. Ca2+ release channels recorded from nondegraded cardiac and skeletal membrane vesicle preparations had slope conductances of 85 and 110 pS, respectively, measured with 1 μM cis-Ca2+ and 50 mM trans-Ba2+. Proteolysis did not alter the unitary channel conductances but did increase the percentage of channel open times from 36{\%} to more than 90{\%}. After proteolysis, channel opening remained dependent on micromolar cis-Ca2+, and high concentrations of ryanodine (300 μM) still blocked the channel. Our results suggest that proteolysis of the Ca2+ release channel with calpain II selectively impairs its inactivation, leaving its unitary conductance and the requirement for micromolar Ca2+ intact.",
keywords = "calcium release channel, calpain, excitation-contraction coupling, sarcoplasmic reticulum",
author = "Rardon, {D. P.} and Cefali, {D. C.} and Mitchell, {R. D.} and Seiler, {S. M.} and Hathaway, {D. R.} and Larry Jones",
year = "1990",
language = "English",
volume = "67",
pages = "84--96",
journal = "Circulation Research",
issn = "0009-7330",
publisher = "Lippincott Williams and Wilkins",
number = "1",

}

TY - JOUR

T1 - Digestion of cardiac and skeletal muscle junctional sarcoplasmic reticulum vesicles with Calpain II. Effects on the Ca2+ release channel

AU - Rardon, D. P.

AU - Cefali, D. C.

AU - Mitchell, R. D.

AU - Seiler, S. M.

AU - Hathaway, D. R.

AU - Jones, Larry

PY - 1990

Y1 - 1990

N2 - The Ca2+ release channel and ryanodine receptor are activities copurifying with the 400,000-450,000 Da high molecular weight protein of cardiac and skeletal junctional sarcoplasmic reticulum. Calpain II, an endogenous cytosolic protease, was used to selectively degrade the high molecular weight protein in cardiac and skeletal muscle sarcoplasmic reticulum vesicles, and its effects on the activity of the Ca2+ release channel and [3H]ryanodine binding sites were analyzed. Degradation of the high molecular weight protein was associated with appearance of 315,000 and 150,000 Da proteolytic fragments and with a change in the ultrastructure of the 'feet', extravesicular projections that protrude from the junctional sarcoplasmic reticulum membrane. The maximal number of [3H]ryanodine binding sites and the affinities of the sites for ryanodine were not remarkably affected by calpain II. Ca2+ release channels recorded from nondegraded cardiac and skeletal membrane vesicle preparations had slope conductances of 85 and 110 pS, respectively, measured with 1 μM cis-Ca2+ and 50 mM trans-Ba2+. Proteolysis did not alter the unitary channel conductances but did increase the percentage of channel open times from 36% to more than 90%. After proteolysis, channel opening remained dependent on micromolar cis-Ca2+, and high concentrations of ryanodine (300 μM) still blocked the channel. Our results suggest that proteolysis of the Ca2+ release channel with calpain II selectively impairs its inactivation, leaving its unitary conductance and the requirement for micromolar Ca2+ intact.

AB - The Ca2+ release channel and ryanodine receptor are activities copurifying with the 400,000-450,000 Da high molecular weight protein of cardiac and skeletal junctional sarcoplasmic reticulum. Calpain II, an endogenous cytosolic protease, was used to selectively degrade the high molecular weight protein in cardiac and skeletal muscle sarcoplasmic reticulum vesicles, and its effects on the activity of the Ca2+ release channel and [3H]ryanodine binding sites were analyzed. Degradation of the high molecular weight protein was associated with appearance of 315,000 and 150,000 Da proteolytic fragments and with a change in the ultrastructure of the 'feet', extravesicular projections that protrude from the junctional sarcoplasmic reticulum membrane. The maximal number of [3H]ryanodine binding sites and the affinities of the sites for ryanodine were not remarkably affected by calpain II. Ca2+ release channels recorded from nondegraded cardiac and skeletal membrane vesicle preparations had slope conductances of 85 and 110 pS, respectively, measured with 1 μM cis-Ca2+ and 50 mM trans-Ba2+. Proteolysis did not alter the unitary channel conductances but did increase the percentage of channel open times from 36% to more than 90%. After proteolysis, channel opening remained dependent on micromolar cis-Ca2+, and high concentrations of ryanodine (300 μM) still blocked the channel. Our results suggest that proteolysis of the Ca2+ release channel with calpain II selectively impairs its inactivation, leaving its unitary conductance and the requirement for micromolar Ca2+ intact.

KW - calcium release channel

KW - calpain

KW - excitation-contraction coupling

KW - sarcoplasmic reticulum

UR - http://www.scopus.com/inward/record.url?scp=0025302280&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0025302280&partnerID=8YFLogxK

M3 - Article

C2 - 2163777

AN - SCOPUS:0025302280

VL - 67

SP - 84

EP - 96

JO - Circulation Research

JF - Circulation Research

SN - 0009-7330

IS - 1

ER -