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. 1985 Dec 1;101(6):2366–2373. doi: 10.1083/jcb.101.6.2366

Sequence and expression of a human type II mesothelial keratin

PMCID: PMC2114014  PMID: 2415537

Abstract

Using mRNA from cultured human mesothelial cells, we constructed bacterial plasmids and lambda phage vectors that contained cDNA sequences specific for the keratins expressed in these cells. A cloned cDNA encoding keratin K7 (55 kD) was identified by positive hybrid selection. Southern Blot analysis indicated that this sequence is represented only once in the human genome, and Northern Blot analysis demonstrated that the gene encoding K7 is expressed in abundance in cultured bronchial and mesothelial cells, but only weakly in cultured epidermal cells and not at all in liver, colon, or exocervical tissue. The predicted amino acid sequence of this keratin has revealed a striking difference between this keratin and the type II keratins expressed in epidermal cells: whereas all of the epidermal type II keratins thus far sequenced have long nonhelical termini rich in glycine and serine, this mesothelial type II keratin has amino and carboxy terminal regions that are unusually short and lack the inexact repeats of glycine and serine residues.

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Selected References

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  1. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  2. Bladon P. T., Bowden P. E., Cunliffe W. J., Wood E. J. Prekeratin biosynthesis in human scalp epidermis. Biochem J. 1982 Oct 15;208(1):179–187. doi: 10.1042/bj2080179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blobel G. A., Moll R., Franke W. W., Vogt-Moykopf I. Cytokeratins in normal lung and lung carcinomas. I. Adenocarcinomas, squamous cell carcinomas and cultured cell lines. Virchows Arch B Cell Pathol Incl Mol Pathol. 1984;45(4):407–429. doi: 10.1007/BF02889883. [DOI] [PubMed] [Google Scholar]
  4. Chang A. C., Nunberg J. H., Kaufman R. J., Erlich H. A., Schimke R. T., Cohen S. N. Phenotypic expression in E. coli of a DNA sequence coding for mouse dihydrofolate reductase. Nature. 1978 Oct 19;275(5681):617–624. doi: 10.1038/275617a0. [DOI] [PubMed] [Google Scholar]
  5. Chou P. Y., Fasman G. D. Prediction of beta-turns. Biophys J. 1979 Jun;26(3):367–383. doi: 10.1016/S0006-3495(79)85259-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chou P. Y., Fasman G. D. Prediction of the secondary structure of proteins from their amino acid sequence. Adv Enzymol Relat Areas Mol Biol. 1978;47:45–148. doi: 10.1002/9780470122921.ch2. [DOI] [PubMed] [Google Scholar]
  7. Cleveland D. W., Lopata M. A., MacDonald R. J., Cowan N. J., Rutter W. J., Kirschner M. W. Number and evolutionary conservation of alpha- and beta-tubulin and cytoplasmic beta- and gamma-actin genes using specific cloned cDNA probes. Cell. 1980 May;20(1):95–105. doi: 10.1016/0092-8674(80)90238-x. [DOI] [PubMed] [Google Scholar]
  8. Connell N. D., Rheinwald J. G. Regulation of the cytoskeleton in mesothelial cells: reversible loss of keratin and increase in vimentin during rapid growth in culture. Cell. 1983 Aug;34(1):245–253. doi: 10.1016/0092-8674(83)90155-1. [DOI] [PubMed] [Google Scholar]
  9. Craig E. A., McCarthy B. J., Wadsworth S. C. Sequence organization of two recombinant plasmids containing genes for the major heat shock-induced protein of D. melanogaster. Cell. 1979 Mar;16(3):575–588. doi: 10.1016/0092-8674(79)90031-x. [DOI] [PubMed] [Google Scholar]
  10. Dowling L. M., Parry D. A., Sparrow L. G. Structural homology between hard alpha-keratin and the intermediate filament proteins desmin and vimentin. Biosci Rep. 1983 Jan;3(1):73–78. doi: 10.1007/BF01121573. [DOI] [PubMed] [Google Scholar]
  11. Eichner R., Bonitz P., Sun T. T. Classification of epidermal keratins according to their immunoreactivity, isoelectric point, and mode of expression. J Cell Biol. 1984 Apr;98(4):1388–1396. doi: 10.1083/jcb.98.4.1388. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fitzgerald M., Shenk T. The sequence 5'-AAUAAA-3'forms parts of the recognition site for polyadenylation of late SV40 mRNAs. Cell. 1981 Apr;24(1):251–260. doi: 10.1016/0092-8674(81)90521-3. [DOI] [PubMed] [Google Scholar]
  13. Franke W. W., Schiller D. L., Hatzfeld M., Winter S. Protein complexes of intermediate-sized filaments: melting of cytokeratin complexes in urea reveals different polypeptide separation characteristics. Proc Natl Acad Sci U S A. 1983 Dec;80(23):7113–7117. doi: 10.1073/pnas.80.23.7113. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fuchs E. V., Coppock S. M., Green H., Cleveland D. W. Two distinct classes of keratin genes and their evolutionary significance. Cell. 1981 Nov;27(1 Pt 2):75–84. doi: 10.1016/0092-8674(81)90362-7. [DOI] [PubMed] [Google Scholar]
  15. Fuchs E., Green H. Changes in keratin gene expression during terminal differentiation of the keratinocyte. Cell. 1980 Apr;19(4):1033–1042. doi: 10.1016/0092-8674(80)90094-x. [DOI] [PubMed] [Google Scholar]
  16. Fuchs E., Hanukoglu I. Unraveling the structure of the intermediate filaments. Cell. 1983 Sep;34(2):332–334. doi: 10.1016/0092-8674(83)90367-7. [DOI] [PubMed] [Google Scholar]
  17. Fuchs E., Marchuk D. Type I and type II keratins have evolved from lower eukaryotes to form the epidermal intermediate filaments in mammalian skin. Proc Natl Acad Sci U S A. 1983 Oct;80(19):5857–5861. doi: 10.1073/pnas.80.19.5857. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Garnier J., Osguthorpe D. J., Robson B. Analysis of the accuracy and implications of simple methods for predicting the secondary structure of globular proteins. J Mol Biol. 1978 Mar 25;120(1):97–120. doi: 10.1016/0022-2836(78)90297-8. [DOI] [PubMed] [Google Scholar]
  19. Girvitz S. C., Bacchetti S., Rainbow A. J., Graham F. L. A rapid and efficient procedure for the purification of DNA from agarose gels. Anal Biochem. 1980 Aug;106(2):492–496. doi: 10.1016/0003-2697(80)90553-9. [DOI] [PubMed] [Google Scholar]
  20. Grunstein M., Hogness D. S. Colony hybridization: a method for the isolation of cloned DNAs that contain a specific gene. Proc Natl Acad Sci U S A. 1975 Oct;72(10):3961–3965. doi: 10.1073/pnas.72.10.3961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Gubler U., Hoffman B. J. A simple and very efficient method for generating cDNA libraries. Gene. 1983 Nov;25(2-3):263–269. doi: 10.1016/0378-1119(83)90230-5. [DOI] [PubMed] [Google Scholar]
  22. Hanukoglu I., Fuchs E. The cDNA sequence of a Type II cytoskeletal keratin reveals constant and variable structural domains among keratins. Cell. 1983 Jul;33(3):915–924. doi: 10.1016/0092-8674(83)90034-x. [DOI] [PubMed] [Google Scholar]
  23. Hanukoglu I., Fuchs E. The cDNA sequence of a human epidermal keratin: divergence of sequence but conservation of structure among intermediate filament proteins. Cell. 1982 Nov;31(1):243–252. doi: 10.1016/0092-8674(82)90424-x. [DOI] [PubMed] [Google Scholar]
  24. Jackson B. W., Grund C., Schmid E., Bürki K., Franke W. W., Illmensee K. Formation of cytoskeletal elements during mouse embryogenesis. Intermediate filaments of the cytokeratin type and desmosomes in preimplantation embryos. Differentiation. 1980;17(3):161–179. doi: 10.1111/j.1432-0436.1980.tb01093.x. [DOI] [PubMed] [Google Scholar]
  25. Johnson L. D., Idler W. W., Zhou X. M., Roop D. R., Steinert P. M. Structure of a gene for the human epidermal 67-kDa keratin. Proc Natl Acad Sci U S A. 1985 Apr;82(7):1896–1900. doi: 10.1073/pnas.82.7.1896. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Jorcano J. L., Franz J. K., Franke W. W. Amino acid sequence diversity between bovine epidermal cytokeratin polypeptides of the basic (type II) subfamily as determined from cDNA clones. Differentiation. 1984;28(2):155–163. doi: 10.1111/j.1432-0436.1984.tb00278.x. [DOI] [PubMed] [Google Scholar]
  27. Jorcano J. L., Magin T. M., Franke W. W. Cell type-specific expression of bovine keratin genes as demonstrated by the use of complementary DNA clones. J Mol Biol. 1984 Jun 15;176(1):21–37. doi: 10.1016/0022-2836(84)90380-2. [DOI] [PubMed] [Google Scholar]
  28. Jorcano J. L., Rieger M., Franz J. K., Schiller D. L., Moll R., Franke W. W. Identification of two types of keratin polypeptides within the acidic cytokeratin subfamily I. J Mol Biol. 1984 Oct 25;179(2):257–281. doi: 10.1016/0022-2836(84)90468-6. [DOI] [PubMed] [Google Scholar]
  29. Kim K. H., Rheinwald J. G., Fuchs E. V. Tissue specificity of epithelial keratins: differential expression of mRNAs from two multigene families. Mol Cell Biol. 1983 Apr;3(4):495–502. doi: 10.1128/mcb.3.4.495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. LaRocca P. J., Rheinwald J. G. Coexpression of simple epithelial keratins and vimentin by human mesothelium and mesothelioma in vivo and in culture. Cancer Res. 1984 Jul;44(7):2991–2999. [PubMed] [Google Scholar]
  31. 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]
  32. Magin T. M., Jorcano J. L., Franke W. W. Translational products of mRNAs coding for non-epidermal cytokeratins. EMBO J. 1983;2(8):1387–1392. doi: 10.1002/j.1460-2075.1983.tb01596.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Marchuk D., McCrohon S., Fuchs E. Complete sequence of a gene encoding a human type I keratin: sequences homologous to enhancer elements in the regulatory region of the gene. Proc Natl Acad Sci U S A. 1985 Mar;82(6):1609–1613. doi: 10.1073/pnas.82.6.1609. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Marchuk D., McCrohon S., Fuchs E. Remarkable conservation of structure among intermediate filament genes. Cell. 1984 Dec;39(3 Pt 2):491–498. doi: 10.1016/0092-8674(84)90456-2. [DOI] [PubMed] [Google Scholar]
  35. McLachlan A. D. Coiled coil formation and sequence regularities in the helical regions of alpha-keratin. J Mol Biol. 1978 Sep 5;124(1):297–304. doi: 10.1016/0022-2836(78)90163-8. [DOI] [PubMed] [Google Scholar]
  36. McLachlan A. D., Stewart M. Tropomyosin coiled-coil interactions: evidence for an unstaggered structure. J Mol Biol. 1975 Oct 25;98(2):293–304. doi: 10.1016/s0022-2836(75)80119-7. [DOI] [PubMed] [Google Scholar]
  37. Moll R., Franke W. W., Schiller D. L., Geiger B., Krepler R. The catalog of human cytokeratins: patterns of expression in normal epithelia, tumors and cultured cells. Cell. 1982 Nov;31(1):11–24. doi: 10.1016/0092-8674(82)90400-7. [DOI] [PubMed] [Google Scholar]
  38. Rheinwald J. G., Green H. Epidermal growth factor and the multiplication of cultured human epidermal keratinocytes. Nature. 1977 Feb 3;265(5593):421–424. doi: 10.1038/265421a0. [DOI] [PubMed] [Google Scholar]
  39. Rheinwald J. G., Green H. Serial cultivation of strains of human epidermal keratinocytes: the formation of keratinizing colonies from single cells. Cell. 1975 Nov;6(3):331–343. doi: 10.1016/s0092-8674(75)80001-8. [DOI] [PubMed] [Google Scholar]
  40. Rheinwald J. G. Serial cultivation of normal human epidermal keratinocytes. Methods Cell Biol. 1980;21A:229–254. doi: 10.1016/s0091-679x(08)60769-4. [DOI] [PubMed] [Google Scholar]
  41. Ricciardi R. P., Miller J. S., Roberts B. E. Purification and mapping of specific mRNAs by hybridization-selection and cell-free translation. Proc Natl Acad Sci U S A. 1979 Oct;76(10):4927–4931. doi: 10.1073/pnas.76.10.4927. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Roop D. R., Hawley-Nelson P., Cheng C. K., Yuspa S. H. Keratin gene expression in mouse epidermis and cultured epidermal cells. Proc Natl Acad Sci U S A. 1983 Feb;80(3):716–720. doi: 10.1073/pnas.80.3.716. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Sanger F., Coulson A. R., Barrell B. G., Smith A. J., Roe B. A. Cloning in single-stranded bacteriophage as an aid to rapid DNA sequencing. J Mol Biol. 1980 Oct 25;143(2):161–178. doi: 10.1016/0022-2836(80)90196-5. [DOI] [PubMed] [Google Scholar]
  44. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  45. Steinert P. M., Idler W. W., Zimmerman S. B. Self-assembly of bovine epidermal keratin filaments in vitro. J Mol Biol. 1976 Dec 15;108(3):547–567. doi: 10.1016/s0022-2836(76)80136-2. [DOI] [PubMed] [Google Scholar]
  46. Steinert P. M., Parry D. A., Racoosin E. L., Idler W. W., Steven A. C., Trus B. L., Roop D. R. The complete cDNA and deduced amino acid sequence of a type II mouse epidermal keratin of 60,000 Da: analysis of sequence differences between type I and type II keratins. Proc Natl Acad Sci U S A. 1984 Sep;81(18):5709–5713. doi: 10.1073/pnas.81.18.5709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Steinert P. M., Rice R. H., Roop D. R., Trus B. L., Steven A. C. Complete amino acid sequence of a mouse epidermal keratin subunit and implications for the structure of intermediate filaments. Nature. 1983 Apr 28;302(5911):794–800. doi: 10.1038/302794a0. [DOI] [PubMed] [Google Scholar]
  48. Sun T. T., Green H. Keratin filaments of cultured human epidermal cells. Formation of intermolecular disulfide bonds during terminal differentiation. J Biol Chem. 1978 Mar 25;253(6):2053–2060. [PubMed] [Google Scholar]
  49. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Tyner A. L., Eichman M. J., Fuchs E. The sequence of a type II keratin gene expressed in human skin: conservation of structure among all intermediate filament genes. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4683–4687. doi: 10.1073/pnas.82.14.4683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Wickens M. P., Buell G. N., Schimke R. T. Synthesis of double-stranded DNA complementary to lysozyme, ovomucoid, and ovalbumin mRNAs. Optimization for full length second strand synthesis by Escherichia coli DNA polymerase I. J Biol Chem. 1978 Apr 10;253(7):2483–2495. [PubMed] [Google Scholar]
  52. Wu Y. J., Parker L. M., Binder N. E., Beckett M. A., Sinard J. H., Griffiths C. T., Rheinwald J. G. The mesothelial keratins: a new family of cytoskeletal proteins identified in cultured mesothelial cells and nonkeratinizing epithelia. Cell. 1982 Dec;31(3 Pt 2):693–703. doi: 10.1016/0092-8674(82)90324-5. [DOI] [PubMed] [Google Scholar]
  53. Young R. A., Davis R. W. Efficient isolation of genes by using antibody probes. Proc Natl Acad Sci U S A. 1983 Mar;80(5):1194–1198. doi: 10.1073/pnas.80.5.1194. [DOI] [PMC free article] [PubMed] [Google Scholar]

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