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. 1985 Oct 1;101(4):1363–1370. doi: 10.1083/jcb.101.4.1363

Preparation of a gap junction fraction from uteri of pregnant rats: the 28-kD polypeptides of uterus, liver, and heart gap junctions are homologous

PMCID: PMC2113926  PMID: 4044640

Abstract

A procedure for the preparation of a gap junction fraction from the uteri of pregnant rats is described. The uterine gap junctions, when examined by electron microscopy of thin sections and in negatively stained preparations, were similar to gap junctions isolated from heart and liver. Major proteins of similar apparent molecular weight (Mr 28,000) were found in gap junction fractions isolated from the uterus, heart, and liver, and were shown to have highly homologous structures by two-dimensional mapping of their tryptic peptides. An Mr 10,000 polypeptide, previously deduced to be a proteolytic product of the Mr 28,000 polypeptide of rat liver (Nicholson, B. J., L. J. Takemoto, M. W. Hunkapiller, L. E. Hood, and J.-P. Revel, 1983, Cell, 32:967-978), was also studied and shown by chymotryptic mapping to be homologous in the uterine, heart, and liver gap junction fractions. An antibody raised in rabbits to a synthetic peptide corresponding to an amino- terminal sequence of the liver gap junction protein recognized Mr 28,000 proteins in the three tissues studied, showing that the proteins shared common antigenic determinants. These results indicate that gap junctions are biochemically conserved plasma membrane specializations. The view that gap junctions are tissue-specific plasma membrane organelles based on previous comparisons of Mr 26,000-30,000 polypeptides is not sustained by the present results.

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Selected References

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  1. Alcalá J., Lieska N., Maisel H. Protein composition of bovine lens cortical fiber cell membranes. Exp Eye Res. 1975 Dec;21(6):581–595. doi: 10.1016/0014-4835(75)90040-8. [DOI] [PubMed] [Google Scholar]
  2. Bok D., Dockstader J., Horwitz J. Immunocytochemical localization of the lens main intrinsic polypeptide (MIP26) in communicating junctions. J Cell Biol. 1982 Jan;92(1):213–220. doi: 10.1083/jcb.92.1.213. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Colaco C. A., Evans W. H. A biochemical dissection of the cardiac intercalated disk: isolation of subcellular fractions containing fascia adherentes and gap junctions. J Cell Sci. 1981 Dec;52:313–325. doi: 10.1242/jcs.52.1.313. [DOI] [PubMed] [Google Scholar]
  4. Dermietzel R., Leibstein A., Frixen U., Janssen-Timmen U., Traub O., Willecke K. Gap junctions in several tissues share antigenic determinants with liver gap junctions. EMBO J. 1984 Oct;3(10):2261–2270. doi: 10.1002/j.1460-2075.1984.tb02124.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Elder J. H., Pickett R. A., 2nd, Hampton J., Lerner R. A. Radioiodination of proteins in single polyacrylamide gel slices. Tryptic peptide analysis of all the major members of complex multicomponent systems using microgram quantities of total protein. J Biol Chem. 1977 Sep 25;252(18):6510–6515. [PubMed] [Google Scholar]
  6. Epstein M. L., Gilula N. B. A study of communication specificity between cells in culture. J Cell Biol. 1977 Dec;75(3):769–787. doi: 10.1083/jcb.75.3.769. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Finbow M. E., Shuttleworth J., Hamilton A. E., Pitts J. D. Analysis of vertebrate gap junction protein. EMBO J. 1983;2(9):1479–1486. doi: 10.1002/j.1460-2075.1983.tb01611.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Finbow M., Yancey S. B., Johnson R., Revel J. P. Independent lines of evidence suggesting a major gap junctional protein with a molecular weight of 26,000. Proc Natl Acad Sci U S A. 1980 Feb;77(2):970–974. doi: 10.1073/pnas.77.2.970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fitzgerald P. G., Bok D., Horwitz J. Immunocytochemical localization of the main intrinsic polypeptide (MIP) in ultrathin frozen sections of rat lens. J Cell Biol. 1983 Nov;97(5 Pt 1):1491–1499. doi: 10.1083/jcb.97.5.1491. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. GREENWOOD F. C., HUNTER W. M., GLOVER J. S. THE PREPARATION OF I-131-LABELLED HUMAN GROWTH HORMONE OF HIGH SPECIFIC RADIOACTIVITY. Biochem J. 1963 Oct;89:114–123. doi: 10.1042/bj0890114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Griepp E. B., Bernfield M. R. Acquisition of synchronous beating between embryonic heart cell aggregates and layers. Exp Cell Res. 1978 May;113(2):263–272. doi: 10.1016/0014-4827(78)90366-x. [DOI] [PubMed] [Google Scholar]
  12. Gros D. B., Nicholson B. J., Revel J. P. Comparative analysis of the gap junction protein from rat heart and liver: is there a tissue specificity of gap junctions? Cell. 1983 Dec;35(2 Pt 1):539–549. doi: 10.1016/0092-8674(83)90188-5. [DOI] [PubMed] [Google Scholar]
  13. Henderson D., Eibl H., Weber K. Structure and biochemistry of mouse hepatic gap junctions. J Mol Biol. 1979 Aug 5;132(2):193–218. doi: 10.1016/0022-2836(79)90391-7. [DOI] [PubMed] [Google Scholar]
  14. Hertzberg E. L. A detergent-independent procedure for the isolation of gap junctions from rat liver. J Biol Chem. 1984 Aug 10;259(15):9936–9943. [PubMed] [Google Scholar]
  15. Hertzberg E. L., Gilula N. B. Isolation and characterization of gap junctions from rat liver. J Biol Chem. 1979 Mar 25;254(6):2138–2147. [PubMed] [Google Scholar]
  16. Hertzberg E. L., Gilula N. B. Liver gap junctions and lens fiber junctions: comparative analysis and calmodulin interaction. Cold Spring Harb Symp Quant Biol. 1982;46(Pt 2):639–645. doi: 10.1101/sqb.1982.046.01.060. [DOI] [PubMed] [Google Scholar]
  17. Hertzberg E. L., Lawrence T. S., Gilula N. B. Gap junctional communication. Annu Rev Physiol. 1981;43:479–491. doi: 10.1146/annurev.ph.43.030181.002403. [DOI] [PubMed] [Google Scholar]
  18. Hertzberg E. L., Skibbens R. V. A protein homologous to the 27,000 dalton liver gap junction protein is present in a wide variety of species and tissues. Cell. 1984 Nov;39(1):61–69. doi: 10.1016/0092-8674(84)90191-0. [DOI] [PubMed] [Google Scholar]
  19. Hirokawa N., Heuser J. The inside and outside of gap-junction membranes visualized by deep etching. Cell. 1982 Sep;30(2):395–406. doi: 10.1016/0092-8674(82)90237-9. [DOI] [PubMed] [Google Scholar]
  20. Hope J., Zervos A., Evans W. H. Molecular approaches to the structure and biogenesis of gap junctions. Prog Clin Biol Res. 1984;151:261–274. [PubMed] [Google Scholar]
  21. Hunter G. K., Pitts J. D. Non-selective junctional communication between some different mammalian cell types in primary culture. J Cell Sci. 1981 Jun;49:163–175. doi: 10.1242/jcs.49.1.163. [DOI] [PubMed] [Google Scholar]
  22. Janssen-Timmen U., Dermietzel R., Frixen U., Leibstein A., Traub O., Willecke K. Immunocytochemical localization of the gap junction 26 K protein in mouse liver plasma membranes. EMBO J. 1983;2(3):295–302. doi: 10.1002/j.1460-2075.1983.tb01422.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kensler R. W., Goodenough D. A. Isolation of mouse myocardial gap junctions. J Cell Biol. 1980 Sep;86(3):755–764. doi: 10.1083/jcb.86.3.755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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]
  25. Loewenstein W. R. Junctional intercellular communication and the control of growth. Biochim Biophys Acta. 1979 Feb 4;560(1):1–65. doi: 10.1016/0304-419x(79)90002-7. [DOI] [PubMed] [Google Scholar]
  26. Manjunath C. K., Goings G. E., Page E. Cytoplasmic surface and intramembrane components of rat heart gap junctional proteins. Am J Physiol. 1984 Jun;246(6 Pt 2):H865–H875. doi: 10.1152/ajpheart.1984.246.6.H865. [DOI] [PubMed] [Google Scholar]
  27. Manjunath C. K., Goings G. E., Page E. Isolation and protein composition of gap junctions from rabbit hearts. Biochem J. 1982 Jul 1;205(1):189–194. doi: 10.1042/bj2050189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Neville D. M., Jr Isolation of an organ specific protein antigen from cell-surface membrane of rat liver. Biochim Biophys Acta. 1968 Apr 9;154(3):540–552. doi: 10.1016/0005-2795(68)90014-7. [DOI] [PubMed] [Google Scholar]
  29. Nicholson B. J., Hunkapiller M. W., Grim L. B., Hood L. E., Revel J. P. Rat liver gap junction protein: properties and partial sequence. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7594–7598. doi: 10.1073/pnas.78.12.7594. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Nicholson B. J., Takemoto L. J., Hunkapiller M. W., Hood L. E., Revel J. P. Differences between liver gap junction protein and lens MIP 26 from rat: implications for tissue specificity of gap junctions. Cell. 1983 Mar;32(3):967–978. doi: 10.1016/0092-8674(83)90081-8. [DOI] [PubMed] [Google Scholar]
  31. Paul D. L., Goodenough D. A. Preparation, characterization, and localization of antisera against bovine MP26, an integral protein from lens fiber plasma membrane. J Cell Biol. 1983 Mar;96(3):625–632. doi: 10.1083/jcb.96.3.625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Pitts J. D., Burk R. R. Specificity of junctional communication between animal cells. Nature. 1976 Dec 23;264(5588):762–764. doi: 10.1038/264762a0. [DOI] [PubMed] [Google Scholar]
  33. Pitts J. D. The role of junctional communication in animal tissues. In Vitro. 1980 Dec;16(12):1049–1056. doi: 10.1007/BF02619255. [DOI] [PubMed] [Google Scholar]
  34. Puri C. P., Garfield R. E. Changes in hormone levels and gap junctions in the rat uterus during pregnancy and parturition. Biol Reprod. 1982 Nov;27(4):967–975. doi: 10.1095/biolreprod27.4.967. [DOI] [PubMed] [Google Scholar]
  35. Revel J. P., Nicholson B. J., Yancey S. B. Partial sequence and turnover of rat liver gap junction protein. Cold Spring Harb Symp Quant Biol. 1982;46(Pt 2):633–637. doi: 10.1101/sqb.1982.046.01.059. [DOI] [PubMed] [Google Scholar]
  36. Sims S. M., Daniel E. E., Garfield R. E. Improved electrical coupling in uterine smooth muscle is associated with increased numbers of gap junctions at parturition. J Gen Physiol. 1982 Sep;80(3):353–375. doi: 10.1085/jgp.80.3.353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Takemoto L. J., Hansen J. S., Nicholson B. J., Hunkapiller M., Revel J. P., Horwitz J. Major intrinsic polypeptide of lens membrane. Biochemical and immunological characterization of the major cyanogen bromide fragment. Biochim Biophys Acta. 1983 Jun 10;731(2):267–274. doi: 10.1016/0005-2736(83)90018-4. [DOI] [PubMed] [Google Scholar]
  38. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Warner A. E., Guthrie S. C., Gilula N. B. Antibodies to gap-junctional protein selectively disrupt junctional communication in the early amphibian embryo. Nature. 1984 Sep 13;311(5982):127–131. doi: 10.1038/311127a0. [DOI] [PubMed] [Google Scholar]
  40. Zampighi G., Simon S. A., Robertson J. D., McIntosh T. J., Costello M. J. On the structural organization of isolated bovine lens fiber junctions. J Cell Biol. 1982 Apr;93(1):175–189. doi: 10.1083/jcb.93.1.175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Zampighi G., Unwin P. N. Two forms of isolated gap junctions. J Mol Biol. 1979 Dec 5;135(2):451–464. doi: 10.1016/0022-2836(79)90446-7. [DOI] [PubMed] [Google Scholar]

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