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. 1985 May 1;100(5):1647–1655. doi: 10.1083/jcb.100.5.1647

Role of myosin in terminal web contraction in isolated intestinal epithelial brush borders

PMCID: PMC2113869  PMID: 3988804

Abstract

We have investigated the role of myosin in contraction of the terminal web in brush borders isolated from intestinal epithelium. At 37 degrees C under conditions that stimulate terminal web contraction (1 microM Ca++ and ATP), most (60-70%) of the myosin is released from the brush border. Approximately 80% of the myosin is also released by ATP at 0 degree C, in the absence of contraction. Preextraction of this 80% of the myosin from brush borders with ATP has no effect on either the time course or extent of subsequently stimulated contraction. However, contraction is inhibited by removal of all of the myosin with 0.6 M KCl and ATP. Contraction is also inhibited by an antibody to brush border myosin, which inhibits both the ATPase activity of brush border myosin and its ability to form stable bipolar polymers. These results indicate that although functional myosin is absolutely required for terminal web contraction only approximately 20% of the brush border myosin is actually necessary. This raises the possibility that there are at least two different subsets of myosin in the terminal web.

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Selected References

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  1. Bentzel C. J., Hainau B., Ho S., Hui S. W., Edelman A., Anagnostopoulos T., Benedetti E. L. Cytoplasmic regulation of tight-junction permeability: effect of plant cytokinins. Am J Physiol. 1980 Sep;239(3):C75–C89. doi: 10.1152/ajpcell.1980.239.3.C75. [DOI] [PubMed] [Google Scholar]
  2. Bretscher A. Microfilament organization in the cytoskeleton of the intestinal brush border. Cell Muscle Motil. 1983;4:239–268. [PubMed] [Google Scholar]
  3. Bretscher A., Weber K. Localization of actin and microfilament-associated proteins in the microvilli and terminal web of the intestinal brush border by immunofluorescence microscopy. J Cell Biol. 1978 Dec;79(3):839–845. doi: 10.1083/jcb.79.3.839. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Broschat K. O., Stidwill R. P., Burgess D. R. Phosphorylation controls brush border motility by regulating myosin structure and association with the cytoskeleton. Cell. 1983 Dec;35(2 Pt 1):561–571. doi: 10.1016/0092-8674(83)90190-3. [DOI] [PubMed] [Google Scholar]
  5. Burgess D. R. Reactivation of intestinal epithelial cell brush border motility: ATP-dependent contraction via a terminal web contractile ring. J Cell Biol. 1982 Dec;95(3):853–863. doi: 10.1083/jcb.95.3.853. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Burgess W. H., Jemiolo D. K., Kretsinger R. H. Interaction of calcium and calmodulin in the presence of sodium dodecyl sulfate. Biochim Biophys Acta. 1980 Jun 26;623(2):257–270. doi: 10.1016/0005-2795(80)90254-8. [DOI] [PubMed] [Google Scholar]
  7. Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
  8. Drenckhahn D., Gröschel-Stewart U. Localization of myosin, actin, and tropomyosin in rat intestinal epithelium: immunohistochemical studies at the light and electron microscope levels. J Cell Biol. 1980 Aug;86(2):475–482. doi: 10.1083/jcb.86.2.475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Duffey M. E., Hainau B., Ho S., Bentzel C. J. Regulation of epithelial tight junction permeability by cyclic AMP. Nature. 1981 Dec 3;294(5840):451–453. doi: 10.1038/294451a0. [DOI] [PubMed] [Google Scholar]
  10. Fairbanks G., Steck T. L., Wallach D. F. Electrophoretic analysis of the major polypeptides of the human erythrocyte membrane. Biochemistry. 1971 Jun 22;10(13):2606–2617. doi: 10.1021/bi00789a030. [DOI] [PubMed] [Google Scholar]
  11. Fechheimer M., Daiss J. L., Cebra J. J. Interaction of immunoglobulin with actin. Mol Immunol. 1979 Nov;16(11):881–888. doi: 10.1016/0161-5890(79)90086-5. [DOI] [PubMed] [Google Scholar]
  12. Fox J. E., Phillips D. R. Role of phosphorylation in mediating the association of myosin with the cytoskeletal structures of human platelets. J Biol Chem. 1982 Apr 25;257(8):4120–4126. [PubMed] [Google Scholar]
  13. Gagelmann M., Rüegg J. C., Di Salvo J. Phosphorylation of the myosin light chains and satellite proteins in detergent-skinned arterial smooth muscle. Biochem Biophys Res Commun. 1984 May 16;120(3):933–938. doi: 10.1016/s0006-291x(84)80196-5. [DOI] [PubMed] [Google Scholar]
  14. Glenney J. R., Jr, Glenney P., Osborn M., Weber K. An F-actin- and calmodulin-binding protein from isolated intestinal brush borders has a morphology related to spectrin. Cell. 1982 Apr;28(4):843–854. doi: 10.1016/0092-8674(82)90063-0. [DOI] [PubMed] [Google Scholar]
  15. Glenney J. R., Jr, Glenney P., Weber K. The spectrin-related molecule, TW-260/240, cross-links the actin bundles of the microvillus rootlets in the brush borders of intestinal epithelial cells. J Cell Biol. 1983 May;96(5):1491–1496. doi: 10.1083/jcb.96.5.1491. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Guerriero V., Jr, Rowley D. R., Means A. R. Production and characterization of an antibody to myosin light chain kinase and intracellular localization of the enzyme. Cell. 1981 Dec;27(3 Pt 2):449–458. doi: 10.1016/0092-8674(81)90386-x. [DOI] [PubMed] [Google Scholar]
  17. Haeberle J. R., Hott J. W., Hathaway D. R. Pseudophosphorylation of the smooth muscle 20 000 dalton myosin light chain. An artifact due to protein modification. Biochim Biophys Acta. 1984 Oct 9;790(1):78–86. doi: 10.1016/0167-4838(84)90334-0. [DOI] [PubMed] [Google Scholar]
  18. Herman I. M., Pollard T. D. Electron microscopic localization of cytoplasmic myosin with ferritin-labeled antibodies. J Cell Biol. 1981 Feb;88(2):346–351. doi: 10.1083/jcb.88.2.346. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hirokawa N., Cheney R. E., Willard M. Location of a protein of the fodrin-spectrin-TW260/240 family in the mouse intestinal brush border. Cell. 1983 Mar;32(3):953–965. doi: 10.1016/0092-8674(83)90080-6. [DOI] [PubMed] [Google Scholar]
  20. Hirokawa N., Keller T. C., 3rd, Chasan R., Mooseker M. S. Mechanism of brush border contractility studied by the quick-freeze, deep-etch method. J Cell Biol. 1983 May;96(5):1325–1336. doi: 10.1083/jcb.96.5.1325. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hirokawa N., Tilney L. G., Fujiwara K., Heuser J. E. Organization of actin, myosin, and intermediate filaments in the brush border of intestinal epithelial cells. J Cell Biol. 1982 Aug;94(2):425–443. doi: 10.1083/jcb.94.2.425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Howe C. L., Mooseker M. S. Characterization of the 110-kdalton actin-calmodulin-, and membrane-binding protein from microvilli of intestinal epithelial cells. J Cell Biol. 1983 Oct;97(4):974–985. doi: 10.1083/jcb.97.4.974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hull B. E., Staehelin L. A. The terminal web. A reevaluation of its structure and function. J Cell Biol. 1979 Apr;81(1):67–82. doi: 10.1083/jcb.81.1.67. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Keller T. C., 3rd, Mooseker M. S. Ca++-calmodulin-dependent phosphorylation of myosin, and its role in brush border contraction in vitro. J Cell Biol. 1982 Dec;95(3):943–959. doi: 10.1083/jcb.95.3.943. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Ledvora R. F., Bárány K., VanderMeulen D. L., Barron J. T., Bárány M. Stretch-induced phosphorylation of the 20,000-dalton light chain of myosin in arterial smooth muscle. J Biol Chem. 1983 Dec 10;258(23):14080–14083. [PubMed] [Google Scholar]
  26. Madara J. L. Increases in guinea pig small intestinal transepithelial resistance induced by osmotic loads are accompanied by rapid alterations in absorptive-cell tight-junction structure. J Cell Biol. 1983 Jul;97(1):125–136. doi: 10.1083/jcb.97.1.125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Mooseker M. S., Bonder E. M., Conzelman K. A., Fishkind D. J., Howe C. L., Keller T. C., 3rd Brush border cytoskeleton and integration of cellular functions. J Cell Biol. 1984 Jul;99(1 Pt 2):104s–112s. doi: 10.1083/jcb.99.1.104s. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mooseker M. S. Brush border motility. Microvillar contraction in triton-treated brush borders isolated from intestinal epithelium. J Cell Biol. 1976 Nov;71(2):417–433. doi: 10.1083/jcb.71.2.417. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Mooseker M. S., Keller T. C., 3rd, Hirokawa N. Regulation of cytoskeletal structure and contractility in the brush border. Ciba Found Symp. 1983;95:195–215. doi: 10.1002/9780470720769.ch12. [DOI] [PubMed] [Google Scholar]
  30. Mooseker M. S., Pollard T. D., Fujiwara K. Characterization and localization of myosin in the brush border of intestinal epithelial cells. J Cell Biol. 1978 Nov;79(2 Pt 1):444–453. doi: 10.1083/jcb.79.2.444. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Nishikawa M., Hidaka H., Adelstein R. S. Phosphorylation of smooth muscle heavy meromyosin by calcium-activated, phospholipid-dependent protein kinase. The effect on actin-activated MgATPase activity. J Biol Chem. 1983 Dec 10;258(23):14069–14072. [PubMed] [Google Scholar]
  32. Owaribe K., Kodama R., Eguchi G. Demonstration of contractility of circumferential actin bundles and its morphogenetic significance in pigmented epithelium in vitro and in vivo. J Cell Biol. 1981 Aug;90(2):507–514. doi: 10.1083/jcb.90.2.507. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Owaribe K., Masuda H. Isolation and characterization of circumferential microfilament bundles from retinal pigmented epithelial cells. J Cell Biol. 1982 Oct;95(1):310–315. doi: 10.1083/jcb.95.1.310. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Pearl M., Fishkind D., Mooseker M., Keene D., Keller T., 3rd Studies on the spectrin-like protein from the intestinal brush border, TW 260/240, and characterization of its interaction with the cytoskeleton and actin. J Cell Biol. 1984 Jan;98(1):66–78. doi: 10.1083/jcb.98.1.66. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Perrie W. T., Perry S. V. An electrophoretic study of the low-molecular-weight components of myosin. Biochem J. 1970 Aug;119(1):31–38. doi: 10.1042/bj1190031. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Pitelka D. R., Taggart B. N. Mechanical tension induces lateral movement of intramembrane components of the tight junction: studies on mouse mammary cells in culture. J Cell Biol. 1983 Mar;96(3):606–612. doi: 10.1083/jcb.96.3.606. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Pollard T. D., Korn E. D. Acanthamoeba myosin. I. Isolation from Acanthamoeba castellanii of an enzyme similar to muscle myosin. J Biol Chem. 1973 Jul 10;248(13):4682–4690. [PubMed] [Google Scholar]
  38. Rodewald R., Newman S. B., Karnovsky M. J. Contraction of isolated brush borders from the intestinal epithelium. J Cell Biol. 1976 Sep;70(3):541–554. doi: 10.1083/jcb.70.3.541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Scholey J. M., Smith R. C., Drenckhahn D., Groschel-Stewart U., Kendrick-Jones J. Thymus myosin. Isolation and characterization of myosin from calf thymus and thymic lymphocytes, and studies on the effect of phosphorylation of its Mr = 20,000 light chain. J Biol Chem. 1982 Jul 10;257(13):7737–7745. [PubMed] [Google Scholar]
  40. Sellers J. R., Eisenberg E., Adelstein R. S. The binding of smooth muscle heavy meromyosin to actin in the presence of ATP. Effect of phosphorylation. J Biol Chem. 1982 Dec 10;257(23):13880–13883. [PubMed] [Google Scholar]
  41. Sheetz M. P., Spudich J. A. Movement of myosin-coated fluorescent beads on actin cables in vitro. Nature. 1983 May 5;303(5912):31–35. doi: 10.1038/303031a0. [DOI] [PubMed] [Google Scholar]

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