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. 1984 Sep 1;99(3):875–885. doi: 10.1083/jcb.99.3.875

Preparation and morphology of sarcoplasmic reticulum terminal cisternae from rabbit skeletal muscle

PMCID: PMC2113387  PMID: 6147356

Abstract

We have developed a procedure to isolate, from skeletal muscle, enriched terminal cisternae of sarcoplasmic reticulum (SR), which retain morphologically intact junctional "feet" structures similar to those observed in situ. The fraction is largely devoid of transverse tubule, plasma membrane, mitochondria, triads (transverse tubules junctionally associated with terminal cisternae), and longitudinal cisternae, as shown by thin-section electron microscopy of representative samples. The terminal cisternae vesicles have distinctive morphological characteristics that differ from the isolated longitudinal cisternae (light SR) obtained from the same gradient. The terminal cisternae consist of two distinct types of membranes, i.e., the junctional face membrane and the Ca2+ pump protein-containing membrane, whereas the longitudinal cisternae contain only the Ca2+ pump protein-containing membrane. The junctional face membrane of the terminal cisternae contains feet structures that extend approximately 12 nm from the membrane surface and can be clearly visualized in thin section through using tannic acid enhancement, by negative staining and by freeze-fracture electron microscopy. Sections of the terminal cisternae, cut tangential to and intersecting the plane of the junctional face, reveal a checkerboardlike lattice of alternating, square-shaped feet structures and spaces each 20 nm square. Structures characteristic of the Ca2+ pump protein are not observed between the feet at the junctional face membrane, either in thin section or by negative staining, even though the Ca2+ pump protein is observed in the nonjunctional membrane on the remainder of the same vesicle. Likewise, freeze-fracture replicas reveal regions of the P face containing ropelike strands instead of the high density of the 7-8-nm particles referable to the Ca2+ pump protein. The intravesicular content of the terminal cisternae, mostly Ca2+-binding protein (calsequestrin), is organized in the form of strands, sometimes appearing paracrystalline, and attached to the inner face of the membrane in the vicinity of the junctional feet. The terminal cisternae preparation is distinct from previously described heavy SR fractions in that it contains the highest percentage of junctional face membrane with morphologically well- preserved junctional feet structures.

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Selected References

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  1. Brunschwig J. P., Brandt N., Caswell A. H., Lukeman D. S. Ultrastructural observations of isolated intact and fragmented junctions of skeletal muscle by use of tannic acid mordanting. J Cell Biol. 1982 Jun;93(3):533–542. doi: 10.1083/jcb.93.3.533. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cadwell J. J., Caswell A. H. Identification of a constituent of the junctional feet linking terminal cisternae to transverse tubules in skeletal muscle. J Cell Biol. 1982 Jun;93(3):543–550. doi: 10.1083/jcb.93.3.543. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Campbell K. P., Franzini-Armstrong C., Shamoo A. E. Further characterization of light and heavy sarcoplasmic reticulum vesicles. Identification of the 'sarcoplasmic reticulum feet' associated with heavy sarcoplasmic reticulum vesicles. Biochim Biophys Acta. 1980 Oct 16;602(1):97–116. doi: 10.1016/0005-2736(80)90293-x. [DOI] [PubMed] [Google Scholar]
  4. Deamer D. W., Baskin R. J. Ultrastructure of sarcoplasmic reticulum preparations. J Cell Biol. 1969 Jul;42(1):296–307. doi: 10.1083/jcb.42.1.296. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Eisenberg B. R., Eisenberg R. S. The T-SR junction in contracting single skeletal muscle fibers. J Gen Physiol. 1982 Jan;79(1):1–19. doi: 10.1085/jgp.79.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Eisenberg B. R., Gilai A. Structural changes in single muscle fibers after stimulation at a low frequency. J Gen Physiol. 1979 Jul;74(1):1–16. doi: 10.1085/jgp.74.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Fleischer S., Fleischer B., Stoeckenius W. Fine structure of lipid-depleted mitochondria. J Cell Biol. 1967 Jan;32(1):193–208. doi: 10.1083/jcb.32.1.193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Franzini-Armstrong C. Membrane particles and transmission at the triad. Fed Proc. 1975 Apr;34(5):1382–1389. [PubMed] [Google Scholar]
  9. Franzini-Armstrong C., Nunzi G. Junctional feet and particles in the triads of a fast-twitch muscle fibre. J Muscle Res Cell Motil. 1983 Apr;4(2):233–252. doi: 10.1007/BF00712033. [DOI] [PubMed] [Google Scholar]
  10. Franzini-Armstrong C. STUDIES OF THE TRIAD : I. Structure of the Junction in Frog Twitch Fibers. J Cell Biol. 1970 Nov 1;47(2):488–499. doi: 10.1083/jcb.47.2.488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Franzini-Armstrong C. Structure of sarcoplasmic reticulum. Fed Proc. 1980 May 15;39(7):2403–2409. [PubMed] [Google Scholar]
  12. Ikemoto N., Nagy B., Bhatnagar G. M., Gergely J. Studies on a metal-binding protein of the sarcoplasmic reticulum. J Biol Chem. 1974 Apr 25;249(8):2357–2365. [PubMed] [Google Scholar]
  13. Jorgensen A. O., Kalnins V., MacLennan D. H. Localization of sarcoplasmic reticulum proteins in rat skeletal muscle by immunofluorescence. J Cell Biol. 1979 Feb;80(2):372–384. doi: 10.1083/jcb.80.2.372. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kelly D. E., Kuda A. M. Subunits of the triadic junction in fast skeletal muscle as revealed by freeze-fracture. J Ultrastruct Res. 1979 Aug;68(2):220–233. doi: 10.1016/s0022-5320(79)90156-4. [DOI] [PubMed] [Google Scholar]
  15. Kelly D. E. The fine structure of skeletal muscle triad junctions. J Ultrastruct Res. 1969 Oct;29(1):37–49. doi: 10.1016/s0022-5320(69)80054-7. [DOI] [PubMed] [Google Scholar]
  16. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. Lau Y. H., Caswell A. H., Brunschwig J. P. Isolation of transverse tubules by fractionation of triad junctions of skeletal muscle. J Biol Chem. 1977 Aug 10;252(15):5565–5574. [PubMed] [Google Scholar]
  19. Meissner G., Conner G. E., Fleischer S. Isolation of sarcoplasmic reticulum by zonal centrifugation and purification of Ca 2+ -pump and Ca 2+ -binding proteins. Biochim Biophys Acta. 1973 Mar 16;298(2):246–269. doi: 10.1016/0005-2736(73)90355-6. [DOI] [PubMed] [Google Scholar]
  20. Meissner G. Isolation and characterization of two types of sarcoplasmic reticulum vesicles. Biochim Biophys Acta. 1975 Apr 21;389(1):51–68. doi: 10.1016/0005-2736(75)90385-5. [DOI] [PubMed] [Google Scholar]
  21. Mitchell R. D., Saito A., Palade P., Fleischer S. Morphology of isolated triads. J Cell Biol. 1983 Apr;96(4):1017–1029. doi: 10.1083/jcb.96.4.1017. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Miyamoto H., Racker E. Calcium-induced calcium release at terminal cisternae of skeletal sarcoplasmic reticulum. FEBS Lett. 1981 Oct 26;133(2):235–238. doi: 10.1016/0014-5793(81)80513-3. [DOI] [PubMed] [Google Scholar]
  23. Ostwald T. J., MacLennan D. H., Dorrington K. J. Effects of cation binding on the conformation of calsequestrin and the high affinity calcium-binding protein of sarcoplasmic reticulum. J Biol Chem. 1974 Sep 25;249(18):5867–5871. [PubMed] [Google Scholar]
  24. Palade P., Saito A., Mitchell R. D., Fleischer S. Preparation of representative samples of subcellular fractions for electron microscopy by filtration with dextran. J Histochem Cytochem. 1983 Jul;31(7):971–974. doi: 10.1177/31.7.6189886. [DOI] [PubMed] [Google Scholar]
  25. Peachey L. D. The sarcoplasmic reticulum and transverse tubules of the frog's sartorius. J Cell Biol. 1965 Jun;25(3 Suppl):209–231. doi: 10.1083/jcb.25.3.209. [DOI] [PubMed] [Google Scholar]
  26. Saito A., Wang C. T., Fleischer S. Membrane asymmetry and enhanced ultrastructural detail of sarcoplasmic reticulum revealed with use of tannic acid. J Cell Biol. 1978 Dec;79(3):601–616. doi: 10.1083/jcb.79.3.601. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Salviati G., Volpe P., Salvatori S., Betto R., Damiani E., Margreth A., Pasquali-Ronchetti I. Biochemical heterogeneity of skeletal-muscle microsomal membranes. Membrane origin, membrane specificity and fibre types. Biochem J. 1982 Feb 15;202(2):289–301. doi: 10.1042/bj2020289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Somlyo A. V. Bridging structures spanning the junctioning gap at the triad of skeletal muscle. J Cell Biol. 1979 Mar;80(3):743–750. doi: 10.1083/jcb.80.3.743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Van Winkle W. B., Bick R. J., Tucker D. E., Tate C. A., Entman M. L. Evidence for membrane microheterogeneity in the sarcoplasmic reticulum of fast twitch skeletal muscle. J Biol Chem. 1982 Oct 10;257(19):11689–11695. [PubMed] [Google Scholar]
  30. Wang C. T., Saito A., Fleischer S. Correlation of ultrastructure of reconstituted sarcoplasmic reticulum membrane vesicles with variation in phospholipid to protein ratio. J Biol Chem. 1979 Sep 25;254(18):9209–9219. [PubMed] [Google Scholar]
  31. Warren G. B., Toon P. A., Birdsall N. J., Lee A. G., Metcalfe J. C. Reconstitution of a calcium pump using defined membrane components. Proc Natl Acad Sci U S A. 1974 Mar;71(3):622–626. doi: 10.1073/pnas.71.3.622. [DOI] [PMC free article] [PubMed] [Google Scholar]

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