Distinct regions of triadin are required for targeting and retention at the junctional domain of the sarcoplasmic reticulum

2014 ◽  
Vol 458 (2) ◽  
pp. 407-417 ◽  
Author(s):  
Daniela Rossi ◽  
Cristina Bencini ◽  
Marina Maritati ◽  
Francesca Benini ◽  
Stefania Lorenzini ◽  
...  
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Stable Association ◽  
Junctional Sarcoplasmic Reticulum

Three regions contribute to triadin localization to the junctional sarcoplasmic reticulum. Dynamics studies revealed that TR3 mediates triadin stability at junctional sites. The stable association of triadin at the junctional sites is facilitated by interactions with calsequestrin-1.

Anchorfibers and the Topography of Junctional SR

1977 ◽  
Vol 35 ◽  
pp. 582-583
Author(s):  
James Junker ◽  
Joachim R. Sommer
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Cardiac Muscle ◽  
Single Filament ◽  
Relaxation Cycle ◽  
10 Nm Filaments ◽  
Junctional Sarcoplasmic Reticulum

Junctional sarcoplasmic reticulum (JSR) in all its forms (extended JSR, JSR of couplings, corbular SR) in both skeletal and cardiac muscle is always located at the Z - I regions of the sarcomeres. The Z tubule is a tubule of the free SR (non-specialized SR) which is consistently located at the Z lines in cardiac muscle (1). Short connections between JSR and Z lines have been described (2), and bundles of filaments at Z lines have been seen in skeletal (3) and cardiac (4) muscle. In opossum cardiac muscle, we have seen bundles of 10 nm filaments stretching across interfibrillary spaces and adjacent myofibrils with extensions to the plasma- lemma in longitudinal (Fig. 1) and transverse (Fig. 2) sections. Only an occasional single filament is seen elsewhere along a sarcomere. We propose that these filaments represent anchor fibers that maintain the observed invariant topography of the free SR and JSR throughout the contraction-relaxation cycle.

Targeting of calsequestrin to sarcoplasmic reticulum after deletions of its acidic carboxy terminus

1999 ◽  
Vol 277 (5) ◽  
pp. C974-C981 ◽  
Author(s):  
Alessandra Nori ◽  
Eleonora Gola ◽  
Stefano Tosato ◽  
Marcello Cantini ◽  
Pompeo Volpe
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Electrostatic Interactions ◽  
Intracellular Localization ◽  
Primary Cultures ◽  
Epifluorescence Microscopy ◽  
Carboxy Terminus ◽  
Adult Rats ◽  
Viral Epitope ◽  
Integral Proteins ◽  
Junctional Sarcoplasmic Reticulum

Calsequestrin (CS) is the Ca2+ binding protein of the junctional sarcoplasmic reticulum (jSR) lumen. Recently, a chimeric CS-HA1, obtained by adding the nine-amino-acid viral epitope hemagglutinin (HA1) to the COOH terminus of CS, was shown to be correctly segregated to the sarcoplasmic reticulum [A. Nori, K. A. Nadalini, A. Martini, R. Rizzuto, A. Villa, and P. Volpe. Am. J. Physiol. 272 ( Cell Physiol. 41): C1420–C1428, 1997]. A putative targeting mechanism of CS to jSR implies electrostatic interactions between negative charges on CS and positive charges on intraluminal domains of jSR integral proteins, such as triadin and junctin. To test this hypothesis, 2 deletion mutants of chimeric CS were engineered: CS-HA1ΔGlu-Asp, in which the 14 acidic residues [-Glu-(Asp)5-Glu-(Asp)7-] of the COOH-terminal tail were removed, and CS-HA1Δ49COOH, in which the last, mostly acidic, 49 residues of the COOH terminus were removed. Both mutant cDNAs were transiently transfected in HeLa cells, myoblasts of rat skeletal muscle primary cultures, or regenerating soleus muscle fibers of adult rats. The expression and intracellular localization of CS-HA1 mutants were studied by epifluorescence microscopy with use of antibodies against CS or HA1. CS-HA1 mutants were shown to be expressed, sorted, and correctly segregated to jSR. Thus short or long deletions of the COOH-terminal acidic tail do not influence the targeting mechanism of CS.

Calcium is released from the junctional sarcoplasmic reticulum during cardiac muscle contraction

1991 ◽  
Vol 260 (3) ◽  
pp. H989-H997 ◽  
Author(s):  
C. S. Moravec ◽  
M. Bond
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Muscle Contraction ◽  
Cardiac Muscle ◽  
Electron Probe Microanalysis ◽  
Electron Probe ◽  
Storage Site ◽  
Papillary Muscles ◽  
Cardiac Muscle Contraction ◽  
Intracellular Storage ◽  
Junctional Sarcoplasmic Reticulum

We have used electron-probe microanalysis (EPMA) to address the question of Ca2+ release by junctional sarcoplasmic reticulum (JSR) as well as Ca2+ regulation by mitochondria (MT) during cardiac muscle contraction. Hamster papillary muscles were rapidly frozen during relaxation or at the peak rate of tension rise (+dT/dt). Total Ca2+ content was measured by EPMA in the JSR, within a MT, over the A band, and in the whole cell, in nine cells per animal (five animals per group). JSR Ca2+ content was found to be significantly lower in muscles frozen at the peak of contraction [7.3 +/- 1.3 (mean +/- SE) mmol Ca2+/kg dry wt] than in those frozen during relaxation (12.5 +/- 1.9 mmol Ca2+/kg dry wt; P less than 0.01), suggesting that Ca2+ is released from this storage site during cardiac muscle contraction. In contrast, MT Ca2+ content did not change significantly during contraction (0.4 +/- 0.1 mmol/kg dry wt) compared with relaxation (0.1 +/- 0.2 mmol/kg dry wt). A third group of muscles was frozen during relaxation after pretreatment with 10(-7) M ryanodine. Ca2+ content of the JSR was significantly decreased (P less than 0.01) in this group of muscles, (6.4 +/- 1.8 mmol/kg dry wt) compared with those frozen during relaxation in the absence of the drug. This suggests that the intracellular storage site with a decreased Ca2+ content in muscles frozen at the peak of contraction is the ryanodine-releasable store. These results provide the first direct measurement of the Ca2+ content of both JSR and MT during a normal cardiac muscle contraction and demonstrate that Ca2+ is released from the JSR during muscle contraction.

X-ray microanalysis of single cardiac myocytes frozen under voltage-clamp conditions

1989 ◽  
Vol 256 (2) ◽  
pp. H574-H583 ◽  
Author(s):  
M. F. Wendt-Gallitelli ◽  
G. Isenberg
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Microprobe Analysis ◽  
Ventricular Myocyte ◽  
Pulse Electron ◽  
Ca Current ◽  
X Ray ◽  
Freeze Dried ◽  
Patch Pipette ◽  
Intracellular Compartments ◽  
Junctional Sarcoplasmic Reticulum

By means of a patch pipette, an isolated ventricular myocyte was transferred into the taper of a silver holder covered by pioloform film. Once the cell was on the film, the cell was voltage clamped (pulses from -45 to +5 mV at 0.5 Hz). The amount of Ca entry was estimated from the Ca current. When contractility (cell shortening) was potentiated with either five pulses of 0.2 s or four pulses of 1 s, shock freezing was timed 116 or 816 ms after start of the clamp pulse. Electron micrographs from freeze-substituted cells revealed the good preservation of the intracellular compartments. The myocytes were cut at -150 degrees C, and the cryosections were freeze dried. In representative examples, the amount of Ca entry is compared with the subcellular Ca distribution as it is analyzed with energy dispersive X-ray microprobe analysis in cytoplasm, junctional sarcoplasmic reticulum (SR), mitochondria, and the subsarcolemmal space (sarcolemma, peripheral SR, fringe of cytosol).

Chimeric calsequestrin and its targeting to the junctional sarcoplasmic reticulum of skeletal muscle

1997 ◽  
Vol 272 (5) ◽  
pp. C1420-C1428 ◽  
Author(s):  
A. Nori ◽  
K. A. Nadalini ◽  
A. Martini ◽  
R. Rizzuto ◽  
A. Villa ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Confocal Microscopy ◽  
Soleus Muscle ◽  
Hela Cells ◽  
Skeletal Muscles ◽  
Intracellular Localization ◽  
Primary Cultures ◽  
Muscle Fibers ◽  
Junctional Sarcoplasmic Reticulum

Calsequestrin (CS) is the junctional sarcoplasmic reticulum (jSR) Ca2+ binding protein responsible for intraluminal Ca2+ storage. The targeting mechanisms of CS to the jSR are yet to be unraveled. The nine-amino acid epitope of the influenza virus hemoagglutinin (referred to as HA1) was added at the COOH-terminal of CS by polymerase chain reaction cloning. The HA1-tagged CS cDNA was transiently transfected in either HeLa cells, myogenic cell lines, such as C2 and L8 cells, myoblasts of rat skeletal muscle primary cultures, or regenerating soleus muscle fibers of adult rats. The expression and intracellular localization of chimeric CS-HA1 were monitored by epifluorescence and confocal microscopy using either anti-CS antibodies or anti-HA1 antibodies. About 30% of transfected HeLa cells and 20-40% of myogenic cells expressed CS-HA1 into intracellular compartments, such as the perinuclear cisternae of endoplasmic reticulum (ER). Myoblasts of newborn rat skeletal muscles were first transfected and subsequently stimulated to differentiate into myotubes. CS-HA1 was detected in approximately 20% of transfected myotubes and did not affect CS distribution in myotubes. In the soleus muscle of adult rat, intramuscular injection of bupivacaine induced necrosis followed by regeneration. In vivo transfection of HA1-tagged CS cDNA in regenerating skeletal muscles determined expression in a few skeletal muscle fibers; CS-HA1 was localized only in jSR, as judged by confocal microscopy of longitudinal sections. The present results show that chimeric CS-HA1 is correctly sorted to ER/SR compartments and that the free COOH-terminal is not requested for sorting, retention, and segregation of CS to the SR.

scholarly journals Cytochemical studies of myocardial adenylate cyclase after its activation and inhibition.

1984 ◽  
Vol 32 (1) ◽  
pp. 105-113 ◽  
Author(s):  
J Slezak ◽  
S A Geller
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Adenylate Cyclase ◽  
Adenosine Deaminase ◽  
Incubation Medium ◽  
Calcium Release ◽  
Inhibitory Effect ◽  
Alkaline Phosphatases ◽  
Similar Role ◽  
Adenosine Triphosphatases ◽  
Junctional Sarcoplasmic Reticulum

Incubation medium, as previously described (J Histochem Cytochem 27:774, 1979), was used to demonstrate the presence of adenylate cyclase (AC) in myocardium. NaF and ouabain were used to inhibit adenosine triphosphatases (ATP) and NaF and isoproterenol were used as activators of AC. The inhibitory effect of adenosine on AC was blocked by the addition of adenosine deaminase. The addition of tetramisol blocked the influence of the alkaline phosphatases on adenylyl imidodiphosphate hydrolysis. The use of these substances resulted in specific precipitation localized in junctional sarcoplasmic reticulum and sarcolemma. The reaction product was dramatically intensified after activation of AC by NaF or isoproterenol. Preincubation in 10-100 mM of propranolol, for 30 min, blocked AC stimulation by isoproterenol and prevented the appearance of the specific precipitate. The localization of specific precipitate in junctional sarcoplasmic reticulum and subsarcolemmal cisternae corresponds to the localization of Na+, K+ ATPase and may reflect the similar role that AC and Na+, K+ ATPase play in calcium release from sarcoplasmic reticulum of internal and peripheral couplings.

scholarly journals SPHEROIDAL BODIES IN THE JUNCTIONAL SARCOPLASMIC RETICULUM OF LIZARD MYOCARDIAL CELLS

1974 ◽  
Vol 60 (3) ◽  
pp. 602-615 ◽  
Author(s):  
M. S. Forbes ◽  
N. Sperelakis
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Anolis Carolinensis ◽  
Myocardial Cells ◽  
En Face ◽  
Structural Details ◽  
Junctional Sarcoplasmic Reticulum

The sarcoplasmic reticulum (SR) of lizard (Anolis carolinensis) myocardial cells has been examined, with particular attention being paid to the structural details of the peripheral couplings (junctional SR). Spheroidal bodies are present within the opaque core of junctional SR; these can be seen both in sections made en face and in sections cut to show the apposition of the junctional SR with the sarcolemma. Opaque junctional processes extend between the sarcolemma and the peripheral junctional SR. The myocardial cells in addition contain some SR cisternae deep within the cells which also possess opaque cores composed of spheroids. Although the significance of the junctional SR spheroidal bodies is unknown, it is thought that they could act as a matrix on which enzymes such as calcium-specific ATPase may be located.

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