Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1994 Feb 15;13(4):744–750. doi: 10.1002/j.1460-2075.1994.tb06316.x

Connexin43 and connexin45 form gap junctions with different molecular permeabilities in osteoblastic cells.

T H Steinberg 1, R Civitelli 1, S T Geist 1, A J Robertson 1, E Hick 1, R D Veenstra 1, H Z Wang 1, P M Warlow 1, E M Westphale 1, J G Laing 1, et al.
PMCID: PMC394870  PMID: 8112289

Abstract

We examined the expression and function of gap junctions in two rat osteoblastic cell lines, ROS 17/2.8 and UMR 106-01. The pattern of expression of gap junction proteins in these two cell lines was distinct: ROS cells expressed only connexin43 on their cell surface, while UMR expressed predominantly connexin45. Immunoprecipitation and RNA blot analysis confirmed the relative quantitation of these connexins. Microinjected ROS cells passed Lucifer yellow to many neighboring cells, but UMR cells were poorly coupled by this criterion. Nevertheless, both UMR and ROS cells were electrically coupled, as characterized by the double whole cell patch-clamp technique. These studies suggested that Cx43 in ROS cells mediated cell-cell coupling for both small ions and larger molecules, but Cx45 in UMR cells allowed passage only of small ions. To demonstrate that the expression of different connexins alone accounted for the lack of dye coupling in UMR cells, we assessed dye coupling in UMR cells transfected with either Cx43 or Cx45. The UMR/Cx43 transfectants were highly dye coupled compared with the untransfected UMR cells, but the UMR/Cx45 transfectants demonstrated no increase in dye transfer. These data demonstrate that different gap junction proteins create channels with different molecular permeabilities; they suggest that different connexins permit different types of signalling between cells.

Full text

PDF

Images in this article

744Image
on p.744
745Image
on p.745
746Image
on p.746
747Image
on p.747
748Image
on p.748

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beyer E. C. Molecular cloning and developmental expression of two chick embryo gap junction proteins. J Biol Chem. 1990 Aug 25;265(24):14439–14443. [PubMed] [Google Scholar]
  2. Beyer E. C., Paul D. L., Goodenough D. A. Connexin family of gap junction proteins. J Membr Biol. 1990 Jul;116(3):187–194. doi: 10.1007/BF01868459. [DOI] [PubMed] [Google Scholar]
  3. Beyer E. C., Paul D. L., Goodenough D. A. Connexin43: a protein from rat heart homologous to a gap junction protein from liver. J Cell Biol. 1987 Dec;105(6 Pt 1):2621–2629. doi: 10.1083/jcb.105.6.2621. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Beyer E. C., Reed K. E., Westphale E. M., Kanter H. L., Larson D. M. Molecular cloning and expression of rat connexin40, a gap junction protein expressed in vascular smooth muscle. J Membr Biol. 1992 Apr;127(1):69–76. doi: 10.1007/BF00232759. [DOI] [PubMed] [Google Scholar]
  5. Beyer E. C., Steinberg T. H. Evidence that the gap junction protein connexin-43 is the ATP-induced pore of mouse macrophages. J Biol Chem. 1991 May 5;266(13):7971–7974. [PubMed] [Google Scholar]
  6. Boland C. J., Fried R. M., Tashjian A. H., Jr Measurement of cytosolic free Ca2+ concentrations in human and rat osteosarcoma cells: actions of bone resorption-stimulating hormones. Endocrinology. 1986 Mar;118(3):980–989. doi: 10.1210/endo-118-3-980. [DOI] [PubMed] [Google Scholar]
  7. Carel J. C., Frazier B., Ley T. J., Holers V. M. Analysis of epitope expression and the functional repertoire of recombinant complement receptor 2 (CR2/CD21) in mouse and human cells. J Immunol. 1989 Aug 1;143(3):923–930. [PubMed] [Google Scholar]
  8. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  9. Civitelli R., Beyer E. C., Warlow P. M., Robertson A. J., Geist S. T., Steinberg T. H. Connexin43 mediates direct intercellular communication in human osteoblastic cell networks. J Clin Invest. 1993 May;91(5):1888–1896. doi: 10.1172/JCI116406. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fishman G. I., Spray D. C., Leinwand L. A. Molecular characterization and functional expression of the human cardiac gap junction channel. J Cell Biol. 1990 Aug;111(2):589–598. doi: 10.1083/jcb.111.2.589. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Flagg-Newton J., Simpson I., Loewenstein W. R. Permeability of the cell-to-cell membrane channels in mammalian cell juncton. Science. 1979 Jul 27;205(4404):404–407. doi: 10.1126/science.377490. [DOI] [PubMed] [Google Scholar]
  12. Fraser J. D., Otawara Y., Price P. A. 1,25-Dihydroxyvitamin D3 stimulates the synthesis of matrix gamma-carboxyglutamic acid protein by osteosarcoma cells. Mutually exclusive expression of vitamin K-dependent bone proteins by clonal osteoblastic cell lines. J Biol Chem. 1988 Jan 15;263(2):911–916. [PubMed] [Google Scholar]
  13. Gimlich R. L., Kumar N. M., Gilula N. B. Differential regulation of the levels of three gap junction mRNAs in Xenopus embryos. J Cell Biol. 1990 Mar;110(3):597–605. doi: 10.1083/jcb.110.3.597. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gunning P., Ponte P., Okayama H., Engel J., Blau H., Kedes L. Isolation and characterization of full-length cDNA clones for human alpha-, beta-, and gamma-actin mRNAs: skeletal but not cytoplasmic actins have an amino-terminal cysteine that is subsequently removed. Mol Cell Biol. 1983 May;3(5):787–795. doi: 10.1128/mcb.3.5.787. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Imanaga I., Kameyama M., Irisawa H. Cell-to-cell diffusion of fluorescent dyes in paired ventricular cells. Am J Physiol. 1987 Jan;252(1 Pt 2):H223–H232. doi: 10.1152/ajpheart.1987.252.1.H223. [DOI] [PubMed] [Google Scholar]
  16. Kanter H. L., Laing J. G., Beyer E. C., Green K. G., Saffitz J. E. Multiple connexins colocalize in canine ventricular myocyte gap junctions. Circ Res. 1993 Aug;73(2):344–350. doi: 10.1161/01.res.73.2.344. [DOI] [PubMed] [Google Scholar]
  17. Kanter H. L., Saffitz J. E., Beyer E. C. Cardiac myocytes express multiple gap junction proteins. Circ Res. 1992 Feb;70(2):438–444. doi: 10.1161/01.res.70.2.438. [DOI] [PubMed] [Google Scholar]
  18. Majeska R. J., Nair B. C., Rodan G. A. Glucocorticoid regulation of alkaline phosphatase in the osteoblastic osteosarcoma cell line ROS 17/2.8. Endocrinology. 1985 Jan;116(1):170–179. doi: 10.1210/endo-116-1-170. [DOI] [PubMed] [Google Scholar]
  19. Majeska R. J., Rodan S. B., Rodan G. A. Parathyroid hormone-responsive clonal cell lines from rat osteosarcoma. Endocrinology. 1980 Nov;107(5):1494–1503. doi: 10.1210/endo-107-5-1494. [DOI] [PubMed] [Google Scholar]
  20. Musil L. S., Beyer E. C., Goodenough D. A. Expression of the gap junction protein connexin43 in embryonic chick lens: molecular cloning, ultrastructural localization, and post-translational phosphorylation. J Membr Biol. 1990 Jun;116(2):163–175. doi: 10.1007/BF01868674. [DOI] [PubMed] [Google Scholar]
  21. Neyton J., Trautmann A. Single-channel currents of an intercellular junction. 1985 Sep 26-Oct 2Nature. 317(6035):331–335. doi: 10.1038/317331a0. [DOI] [PubMed] [Google Scholar]
  22. Nishi M., Kumar N. M., Gilula N. B. Developmental regulation of gap junction gene expression during mouse embryonic development. Dev Biol. 1991 Jul;146(1):117–130. doi: 10.1016/0012-1606(91)90452-9. [DOI] [PubMed] [Google Scholar]
  23. Paul D. L., Ebihara L., Takemoto L. J., Swenson K. I., Goodenough D. A. Connexin46, a novel lens gap junction protein, induces voltage-gated currents in nonjunctional plasma membrane of Xenopus oocytes. J Cell Biol. 1991 Nov;115(4):1077–1089. doi: 10.1083/jcb.115.4.1077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Paul D. L. Molecular cloning of cDNA for rat liver gap junction protein. J Cell Biol. 1986 Jul;103(1):123–134. doi: 10.1083/jcb.103.1.123. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Reed K. E., Westphale E. M., Larson D. M., Wang H. Z., Veenstra R. D., Beyer E. C. Molecular cloning and functional expression of human connexin37, an endothelial cell gap junction protein. J Clin Invest. 1993 Mar;91(3):997–1004. doi: 10.1172/JCI116321. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Rup D. M., Veenstra R. D., Wang H. Z., Brink P. R., Beyer E. C. Chick connexin-56, a novel lens gap junction protein. Molecular cloning and functional expression. J Biol Chem. 1993 Jan 5;268(1):706–712. [PubMed] [Google Scholar]
  27. Shupnik M. A., Tashjian A. H., Jr Epidermal growth factor and phorbol ester actions on human osteosarcoma cells. Characterization of response and nonresponsive cell lines. J Biol Chem. 1982 Oct 25;257(20):12161–12164. [PubMed] [Google Scholar]
  28. Sáez J. C., Connor J. A., Spray D. C., Bennett M. V. Hepatocyte gap junctions are permeable to the second messenger, inositol 1,4,5-trisphosphate, and to calcium ions. Proc Natl Acad Sci U S A. 1989 Apr;86(8):2708–2712. doi: 10.1073/pnas.86.8.2708. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Tenbroek E., Arneson M., Jarvis L., Louis C. The distribution of the fiber cell intrinsic membrane proteins MP20 and connexin46 in the bovine lens. J Cell Sci. 1992 Sep;103(Pt 1):245–257. doi: 10.1242/jcs.103.1.245. [DOI] [PubMed] [Google Scholar]
  30. Veenstra R. D. Voltage-dependent gating of gap junction channels in embryonic chick ventricular cell pairs. Am J Physiol. 1990 Apr;258(4 Pt 1):C662–C672. doi: 10.1152/ajpcell.1990.258.4.C662. [DOI] [PubMed] [Google Scholar]
  31. Veenstra R. D., Wang H. Z., Westphale E. M., Beyer E. C. Multiple connexins confer distinct regulatory and conductance properties of gap junctions in developing heart. Circ Res. 1992 Nov;71(5):1277–1283. doi: 10.1161/01.res.71.5.1277. [DOI] [PubMed] [Google Scholar]
  32. Zhang J. T., Nicholson B. J. Sequence and tissue distribution of a second protein of hepatic gap junctions, Cx26, as deduced from its cDNA. J Cell Biol. 1989 Dec;109(6 Pt 2):3391–3401. doi: 10.1083/jcb.109.6.3391. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES