scholarly journals Identification of a constituent of the junctional feet linking terminal cisternae to transverse tubules in skeletal muscle.

1982 ◽  
Vol 93 (3) ◽  
pp. 543-550 ◽  
Author(s):  
J J Cadwell ◽  
A H Caswell
Keyword(s):  
Skeletal Muscle ◽  
Protein Pair ◽  
Specific Marker ◽  
Molecular Weights ◽  
Single Protein ◽  
T Tubules ◽  
Triad Junction ◽  
Terminal Cisternae ◽  
Substantial Degree ◽  
Triad Junctions

This study describes the biochemical composition of junctional feet in skeletal muscle utilizing a fraction of isolated triad junctions. [3H]Ouabain entrapment was employed as a specific marker for T-tubules. The integrity of the triad junction was assayed by the isopycnic density of [3H]ouabain activity (24-30% sucrose for free T-tubules, 38-42% sucrose for intact triads). Trypsin, chymotrypsin, and pronase all caused separation of T-tubules from terminal cisternae, indicating that the junction is composed as least in part of protein. Trypsin and chymotrypsin hydrolyzed four proteins: the Ca2+ pump, a doublet 325,000, 300,000, and an 80,000 Mr protein. T-tubules which had been labeled covalently with 125I were joined to unlabeled terminal cisternae by treatment with K cacodylate. The reformed triads were separated from free T-tubules and then severed by passage through a French press. When terminal cisternae were separated from T-tubules, some 125I label was transferred from the labeled T-tubules to the unlabeled terminal cisternae. Gel electrophoresis showed that, although T-tubules were originally labeled in a large number of different proteins, only a single protein doublet was significantly labeled in the originally unlabeled terminal cisternae. This protein pair had molecular weights of 325,000 and 300,000 daltons. Transfer of label did not occur to a substantial degree without K cacodylate treatment. We propose that the transfer of 125I label from T-tubules to terminal cisternae during reformation and breakage of the triad junction is a property of the protein which spans the gap between T-tubules and terminal cisternae.

scholarly journals Deficiency of triad junction and contraction in mutant skeletal muscle lacking junctophilin type 1

2001 ◽  
Vol 154 (5) ◽  
pp. 1059-1068 ◽  
Author(s):  
Koichi Ito ◽  
Shinji Komazaki ◽  
Kazushige Sasamoto ◽  
Morikatsu Yoshida ◽  
Miyuki Nishi ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Plasma Membrane ◽  
Physiological Role ◽  
Mutant Mice ◽  
Functional Coupling ◽  
Release Channel ◽  
Signal Conversion ◽  
Triad Junction ◽  
Triad Junctions

In skeletal muscle excitation–contraction (E–C) coupling, the depolarization signal is converted from the intracellular Ca2+ store into Ca2+ release by functional coupling between the cell surface voltage sensor and the Ca2+ release channel on the sarcoplasmic reticulum (SR). The signal conversion occurs in the junctional membrane complex known as the triad junction, where the invaginated plasma membrane called the transverse-tubule (T-tubule) is pinched from both sides by SR membranes. Previous studies have suggested that junctophilins (JPs) contribute to the formation of the junctional membrane complexes by spanning the intracellular store membrane and interacting with the plasma membrane (PM) in excitable cells. Of the three JP subtypes, both type 1 (JP-1) and type 2 (JP-2) are abundantly expressed in skeletal muscle. To examine the physiological role of JP-1 in skeletal muscle, we generated mutant mice lacking JP-1. The JP-1 knockout mice showed no milk suckling and died shortly after birth. Ultrastructural analysis demonstrated that triad junctions were reduced in number, and that the SR was often structurally abnormal in the skeletal muscles of the mutant mice. The mutant muscle developed less contractile force (evoked by low-frequency electrical stimuli) and showed abnormal sensitivities to extracellular Ca2+. Our results indicate that JP-1 contributes to the construction of triad junctions and that it is essential for the efficiency of signal conversion during E–C coupling in skeletal muscle.

A transmission delay and the effect of temperature at the triadic junction of skeletal muscle

1986 ◽  
Vol 229 (1254) ◽  
pp. 97-110 ◽  
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Effect Of Temperature ◽  
Muscle Fibres ◽  
Time Interval ◽  
Strong Temperature Dependence ◽  
Ca Channels ◽  
T Tubules ◽  
Skeletal Muscle Fibres ◽  
Terminal Cisternae

The coupling process at the triadic junctions in skeletal muscle fibres is characterized by a significant latency between the depolarization of the transverse tubular membrane and the release of Ca from the sarcoplasmic reticulum. This time interval, the triadic delay, is sufficiently long to allow for the participation of a chemical process. The strong temperature dependence of the triadic delay ( Q 10 near 2.7) suggests that a sequence of chemical steps may link the electical signal in the T-tubules to the opening of Ca channels in the terminal cisternae of the sarcoplasmic reticulum.

scholarly journals Molecular organization of transverse tubule/sarcoplasmic reticulum junctions during development of excitation-contraction coupling in skeletal muscle.

1994 ◽  
Vol 5 (10) ◽  
pp. 1105-1118 ◽  
Author(s):  
B E Flucher ◽  
S B Andrews ◽  
M P Daniels
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Calcium Release ◽  
Structural Characteristics ◽  
Molecular Organization ◽  
Muscle Action Potential ◽  
T Tubules ◽  
Triad Junction ◽  
First Contact ◽  
The Relationship

The relationship between the molecular composition and organization of the triad junction and the development of excitation-contraction (E-C) coupling was investigated in cultured skeletal muscle. Action potential-induced calcium transients develop concomitantly with the first expression of the dihydropyridine receptor (DHPR) and the ryanodine receptor (RyR), which are colocalized in clusters from the time of their earliest appearance. These DHPR/RyR clusters correspond to junctional domains of the transverse tubules (T-tubules) and sarcoplasmic reticulum (SR), respectively. Thus, at first contact T-tubules and SR form molecularly and structurally specialized membrane domains that support E-C coupling. The earliest T-tubule/SR junctions show structural characteristics of mature triads but are diverse in conformation and typically are formed before the extensive development of myofibrils. Whereas the initial formation of T-tubule/SR junctions is independent of association with myofibrils, the reorganization into proper triads occurs as junctions become associated with the border between the A band and the I band of the sarcomere. This final step in triad formation manifests itself in an increased density and uniformity of junctions in the cytoplasm, which in turn results in increased calcium release and reuptake rates.

scholarly journals Cell fractionation studies indicate that dystrophin is a protein of surface membranes of skeletal muscle

1989 ◽  
Vol 258 (3) ◽  
pp. 837-841 ◽  
Author(s):  
G Salviati ◽  
R Betto ◽  
S Ceoldo ◽  
E Biasia ◽  
E Bonilla ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Cholesterol Content ◽  
Binding Activity ◽  
Cell Fractionation ◽  
Ouabain Binding ◽  
Low Concentrations ◽  
Peripheral Membrane Protein ◽  
T Tubules ◽  
Triton X ◽  
Terminal Cisternae

We studied the subcellular localization of dystrophin in rabbit skeletal muscle. In Western-blot analysis of membrane preparations, dystrophin was associated with the sarcolemmal fraction, as indicated by cholesterol content and co-purification with ouabain-binding activity and beta-adrenergic receptor. Dystrophin was also found with junctional T-tubules, but not with ‘free’ T-tubules, longitudinal portions or terminal cisternae of the sarcoplasmic reticulum. Dystrophin was not solubilized by high salt solutions, but it was solubilized by low concentrations of detergents (Triton X-100 and deoxycholate), suggesting that it is a peripheral membrane protein.

Cryo-Electron Tomography of Isolated Triad Junctions from Skeletal Muscle

2001 ◽  
Vol 7 (S2) ◽  
pp. 94-95 ◽  
Author(s):  
C.-E. Hsieh ◽  
M. Marko ◽  
B.K. Rath ◽  
S. Fleischer ◽  
T. Wagenknecht
Keyword(s):  
Skeletal Muscle ◽  
Plasma Membrane ◽  
Electron Tomography ◽  
Mass Density ◽  
Tomographic Reconstruction ◽  
Three Dimensional ◽  
Cryo Electron Tomography ◽  
Junctional Complexes ◽  
Triad Junction ◽  
Triad Junctions

In skeletal muscle, depolarization of the plasma membrane, which is initiated at the neuromuscular junction, is transduced to a rise in cytoplasmic calcium at specialized structures known as triad junctions (TJs). TJs occur in the myofiber’s interior at regions near the z-lines, where transversely oriented tubular invaginations of the plasma membrane (T-tubules) form junctions with two elements of the sarcoplasmic reticulum (SR). Isolation of membrane fractions that are enriched in junctional complexes and which retain function has been reported.Figure 1 shows a region of an electron micrograph containing an isolated TJ in the frozen-hydrated state. in the orientation shown, two SR-derived vesicles sandwich a flattened vesicle derived from the T-tubule. The junctional regions contain a complex distribution of density, presumably due to proteins that are known to be present in TJs. Electron tomography offers the means to determine the three-dimensional mass density from such micrographs, which would greatly aid in their interpretation. Only recently has the automated data collection technology for determining tomograms of non-stained, frozen-hydrated specimens become available. Here we describe the first tomographic reconstruction of a frozen-hydrated triad junction by automated electron tomography.

scholarly journals Dihydropyridine receptor alpha subunits in normal and dysgenic muscle in vitro: expression of alpha 1 is required for proper targeting and distribution of alpha 2.

1991 ◽  
Vol 115 (5) ◽  
pp. 1345-1356 ◽  
Author(s):  
B E Flucher ◽  
J L Phillips ◽  
J A Powell
Keyword(s):  
Muscle Cells ◽  
Distribution Patterns ◽  
Receptor Alpha ◽  
Alpha Subunits ◽  
Nuclear Domain ◽  
T Tubules ◽  
Immunofluorescence Labeling ◽  
T System ◽  
Triad Junctions

We have studied the subcellular distribution of the alpha 1 and alpha 2 subunits of the skeletal muscle dihydropyridine (DHP) receptor with immunofluorescence labeling of normal and dysgenic (mdg) muscle in culture. In normal myotubes both alpha subunits were localized in clusters associated with the T-tubule membranes of longitudinally as well as transversely oriented T-tubules. The DHP receptor-rich domains may represent the sites where triad junctions with the sarcoplasmic reticulum are being formed. In cultures from dysgenic muscle the alpha 1 subunit was undetectable and the distribution patterns of the alpha 2 subunit were abnormal. The alpha subunit did not form clusters nor was it discretely localized in the T-tubule system. Instead, alpha 2 was found diffusely distributed in parts of the T-system, in structures in the perinuclear region and in the plasma membrane. These results suggest that an interaction between the two alpha subunits is required for the normal distribution of the alpha 2 subunit in the T-tubule membranes. Spontaneous fusion of normal non-muscle cells with dysgenic myotubes resulted in a regional expression of the alpha 1 polypeptide near the foreign nuclei, thus defining the nuclear domain of a T-tubule membrane protein in multi-nucleated muscle cells. Furthermore, the normal intracellular distribution of the alpha 2 polypeptide was restored in domains containing a foreign "rescue" nucleus; this supports the idea that direct interactions between the DHP receptor alpha 1 and alpha 2 subunits are involved in the organization of the junctional T-tubule membranes.

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