Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1990 Feb;10(2):662–671. doi: 10.1128/mcb.10.2.662

Cell-type-specific expression of alternatively spliced human fibronectin IIICS mRNAs.

R P Hershberger 1, L A Culp 1
PMCID: PMC360864  PMID: 1688996

Abstract

Fibronectin polypeptide diversity is generated to a large extent by alternative splicing of the fibronectin primary transcript at three sites: two extra domain exons encoding extra structural repeats and a region of nonhomologous sequence termed the type-III connecting segment (IIICS). A novel double primer extension assay was developed to identify and quantify simultaneously each of the five human IIICS mRNA splicing variants. Expression of the five IIICS variants was analyzed in a variety of human normal and tumor cell types as well as in human liver. Differences in IIICS expression patterns were observed among different cell types, among fibroblasts of different tissue origins, and between comparable normal and transformed cells. The most predominant cell-type-specific differences were in the abundance of the one IIICS- mRNA variant relative to the four IIICS+ variants. The percentage of O variant (IIICS-) mRNAs within the total fibronectin mRNA pool varied between 3 and 17% among tumor cells and between 7 and 46% among normal cells. The O variant composed 57% of the fibronectin mRNA in liver tissue, correlating with the previously described increased abundance of IIICS- polypeptide subunits in plasma fibronectin, compared with those in cellular fibronectins. Additional cell-type-specific changes among the expression levels of the four IIICS+ mRNA variants are consistent with a proposed model in which regulation of an alternative selection of a 3'splice site predominates over regulation of the selection of a 5' splice site in generating specific patterns of IIICS mRNA expression.

Full text

PDF
662

Images in this article

665Image
on p.665
666Image
on p.666

Selected References

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

  1. Akiyama S. K., Yamada K. M. Fibronectin. Adv Enzymol Relat Areas Mol Biol. 1987;59:1–57. doi: 10.1002/9780470123058.ch1. [DOI] [PubMed] [Google Scholar]
  2. Andreadis A., Gallego M. E., Nadal-Ginard B. Generation of protein isoform diversity by alternative splicing: mechanistic and biological implications. Annu Rev Cell Biol. 1987;3:207–242. doi: 10.1146/annurev.cb.03.110187.001231. [DOI] [PubMed] [Google Scholar]
  3. Barone M. V., Henchcliffe C., Baralle F. E., Paolella G. Cell type specific trans-acting factors are involved in alternative splicing of human fibronectin pre-mRNA. EMBO J. 1989 Apr;8(4):1079–1085. doi: 10.1002/j.1460-2075.1989.tb03476.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bernard M. P., Kolbe M., Weil D., Chu M. L. Human cellular fibronectin: comparison of the carboxyl-terminal portion with rat identifies primary structural domains separated by hypervariable regions. Biochemistry. 1985 May 21;24(11):2698–2704. doi: 10.1021/bi00332a016. [DOI] [PubMed] [Google Scholar]
  5. Beyth R. J., Culp L. A. Glycosaminoglycan distribution in substratum adhesion sites of aging human skin fibroblasts, including papillary and reticular subpopulations. Mech Ageing Dev. 1985 Feb;29(2):151–169. doi: 10.1016/0047-6374(85)90015-6. [DOI] [PubMed] [Google Scholar]
  6. Borsi L., Carnemolla B., Castellani P., Rosellini C., Vecchio D., Allemanni G., Chang S. E., Taylor-Papadimitriou J., Pande H., Zardi L. Monoclonal antibodies in the analysis of fibronectin isoforms generated by alternative splicing of mRNA precursors in normal and transformed human cells. J Cell Biol. 1987 Mar;104(3):595–600. doi: 10.1083/jcb.104.3.595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Breitbart R. E., Nadal-Ginard B. Developmentally induced, muscle-specific trans factors control the differential splicing of alternative and constitutive troponin T exons. Cell. 1987 Jun 19;49(6):793–803. doi: 10.1016/0092-8674(87)90617-9. [DOI] [PubMed] [Google Scholar]
  8. Carnemolla B., Balza E., Siri A., Zardi L., Nicotra M. R., Bigotti A., Natali P. G. A tumor-associated fibronectin isoform generated by alternative splicing of messenger RNA precursors. J Cell Biol. 1989 Mar;108(3):1139–1148. doi: 10.1083/jcb.108.3.1139. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Castellani P., Siri A., Rosellini C., Infusini E., Borsi L., Zardi L. Transformed human cells release different fibronectin variants than do normal cells. J Cell Biol. 1986 Nov;103(5):1671–1677. doi: 10.1083/jcb.103.5.1671. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Chabot B., Black D. L., LeMaster D. M., Steitz J. A. The 3' splice site of pre-messenger RNA is recognized by a small nuclear ribonucleoprotein. Science. 1985 Dec 20;230(4732):1344–1349. doi: 10.1126/science.2933810. [DOI] [PubMed] [Google Scholar]
  11. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  12. Dean D. C., Newby R. F., Bourgeois S. Regulation of fibronectin biosynthesis by dexamethasone, transforming growth factor beta, and cAMP in human cell lines. J Cell Biol. 1988 Jun;106(6):2159–2170. doi: 10.1083/jcb.106.6.2159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dufour S., Duband J. L., Humphries M. J., Obara M., Yamada K. M., Thiery J. P. Attachment, spreading and locomotion of avian neural crest cells are mediated by multiple adhesion sites on fibronectin molecules. EMBO J. 1988 Sep;7(9):2661–2671. doi: 10.1002/j.1460-2075.1988.tb03119.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Frendewey D., Keller W. Stepwise assembly of a pre-mRNA splicing complex requires U-snRNPs and specific intron sequences. Cell. 1985 Aug;42(1):355–367. doi: 10.1016/s0092-8674(85)80131-8. [DOI] [PubMed] [Google Scholar]
  15. Garcia-Pardo A., Rostagno A., Frangione B. Primary structure of human plasma fibronectin. Characterization of a 38 kDa domain containing the C-terminal heparin-binding site (Hep III site) and a region of molecular heterogeneity. Biochem J. 1987 Feb 1;241(3):923–928. doi: 10.1042/bj2410923. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Green M. R. Pre-mRNA splicing. Annu Rev Genet. 1986;20:671–708. doi: 10.1146/annurev.ge.20.120186.003323. [DOI] [PubMed] [Google Scholar]
  17. Gutman A., Kornblihtt A. R. Identification of a third region of cell-specific alternative splicing in human fibronectin mRNA. Proc Natl Acad Sci U S A. 1987 Oct;84(20):7179–7182. doi: 10.1073/pnas.84.20.7179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hayashi M., Yamada K. M. Differences in domain structures between plasma and cellular fibronectins. J Biol Chem. 1981 Nov 10;256(21):11292–11300. [PubMed] [Google Scholar]
  19. Humphries M. J., Akiyama S. K., Komoriya A., Olden K., Yamada K. M. Identification of an alternatively spliced site in human plasma fibronectin that mediates cell type-specific adhesion. J Cell Biol. 1986 Dec;103(6 Pt 2):2637–2647. doi: 10.1083/jcb.103.6.2637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Humphries M. J., Akiyama S. K., Komoriya A., Olden K., Yamada K. M. Neurite extension of chicken peripheral nervous system neurons on fibronectin: relative importance of specific adhesion sites in the central cell-binding domain and the alternatively spliced type III connecting segment. J Cell Biol. 1988 Apr;106(4):1289–1297. doi: 10.1083/jcb.106.4.1289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Humphries M. J., Komoriya A., Akiyama S. K., Olden K., Yamada K. M. Identification of two distinct regions of the type III connecting segment of human plasma fibronectin that promote cell type-specific adhesion. J Biol Chem. 1987 May 15;262(14):6886–6892. [PubMed] [Google Scholar]
  22. Hynes R. O. Integrins: a family of cell surface receptors. Cell. 1987 Feb 27;48(4):549–554. doi: 10.1016/0092-8674(87)90233-9. [DOI] [PubMed] [Google Scholar]
  23. Hynes R. Molecular biology of fibronectin. Annu Rev Cell Biol. 1985;1:67–90. doi: 10.1146/annurev.cb.01.110185.000435. [DOI] [PubMed] [Google Scholar]
  24. Kornblihtt A. R., Umezawa K., Vibe-Pedersen K., Baralle F. E. Primary structure of human fibronectin: differential splicing may generate at least 10 polypeptides from a single gene. EMBO J. 1985 Jul;4(7):1755–1759. doi: 10.1002/j.1460-2075.1985.tb03847.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kornblihtt A. R., Vibe-Pedersen K., Baralle F. E. Human fibronectin: cell specific alternative mRNA splicing generates polypeptide chains differing in the number of internal repeats. Nucleic Acids Res. 1984 Jul 25;12(14):5853–5868. doi: 10.1093/nar/12.14.5853. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kornblihtt A. R., Vibe-Pedersen K., Baralle F. E. Human fibronectin: molecular cloning evidence for two mRNA species differing by an internal segment coding for a structural domain. EMBO J. 1984 Jan;3(1):221–226. doi: 10.1002/j.1460-2075.1984.tb01787.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Leff S. E., Evans R. M., Rosenfeld M. G. Splice commitment dictates neuron-specific alternative RNA processing in calcitonin/CGRP gene expression. Cell. 1987 Feb 13;48(3):517–524. doi: 10.1016/0092-8674(87)90202-9. [DOI] [PubMed] [Google Scholar]
  28. Maniatis T., Reed R. The role of small nuclear ribonucleoprotein particles in pre-mRNA splicing. Nature. 1987 Feb 19;325(6106):673–678. doi: 10.1038/325673a0. [DOI] [PubMed] [Google Scholar]
  29. Norton P. A., Hynes R. O. Alternative splicing of chicken fibronectin in embryos and in normal and transformed cells. Mol Cell Biol. 1987 Dec;7(12):4297–4307. doi: 10.1128/mcb.7.12.4297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Oyama F., Murata Y., Suganuma N., Kimura T., Titani K., Sekiguchi K. Patterns of alternative splicing of fibronectin pre-mRNA in human adult and fetal tissues. Biochemistry. 1989 Feb 7;28(3):1428–1434. doi: 10.1021/bi00429a072. [DOI] [PubMed] [Google Scholar]
  31. Padgett R. A., Grabowski P. J., Konarska M. M., Seiler S., Sharp P. A. Splicing of messenger RNA precursors. Annu Rev Biochem. 1986;55:1119–1150. doi: 10.1146/annurev.bi.55.070186.005351. [DOI] [PubMed] [Google Scholar]
  32. Paul J. I., Hynes R. O. Multiple fibronectin subunits and their post-translational modifications. J Biol Chem. 1984 Nov 10;259(21):13477–13487. [PubMed] [Google Scholar]
  33. Paul J. I., Schwarzbauer J. E., Tamkun J. W., Hynes R. O. Cell-type-specific fibronectin subunits generated by alternative splicing. J Biol Chem. 1986 Sep 15;261(26):12258–12265. [PubMed] [Google Scholar]
  34. Petersen T. E., Thøgersen H. C., Skorstengaard K., Vibe-Pedersen K., Sahl P., Sottrup-Jensen L., Magnusson S. Partial primary structure of bovine plasma fibronectin: three types of internal homology. Proc Natl Acad Sci U S A. 1983 Jan;80(1):137–141. doi: 10.1073/pnas.80.1.137. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Ruoslahti E. Fibronectin and its receptors. Annu Rev Biochem. 1988;57:375–413. doi: 10.1146/annurev.bi.57.070188.002111. [DOI] [PubMed] [Google Scholar]
  36. Schafer I. A., Shapiro A., Kovach M., Lang C., Fratianne R. B. The interaction of human papillary and reticular fibroblasts and human keratinocytes in the contraction of three-dimensional floating collagen lattices. Exp Cell Res. 1989 Jul;183(1):112–125. doi: 10.1016/0014-4827(89)90422-9. [DOI] [PubMed] [Google Scholar]
  37. Schwarzbauer J. E., Tamkun J. W., Lemischka I. R., Hynes R. O. Three different fibronectin mRNAs arise by alternative splicing within the coding region. Cell. 1983 Dec;35(2 Pt 1):421–431. doi: 10.1016/0092-8674(83)90175-7. [DOI] [PubMed] [Google Scholar]
  38. Sekiguchi K., Klos A. M., Kurachi K., Yoshitake S., Hakomori S. Human liver fibronectin complementary DNAs: identification of two different messenger RNAs possibly encoding the alpha and beta subunits of plasma fibronectin. Biochemistry. 1986 Aug 26;25(17):4936–4941. doi: 10.1021/bi00365a032. [DOI] [PubMed] [Google Scholar]
  39. Sekiguchi K., Siri A., Zardi L., Hakomori S. Differences in domain structure between human fibronectins isolated from plasma and from culture supernatants of normal and transformed fibroblasts. Studies with domain-specific antibodies. J Biol Chem. 1985 Apr 25;260(8):5105–5114. [PubMed] [Google Scholar]
  40. Sekiguchi K., Titani K. Probing molecular polymorphism of fibronectins with antibodies directed to the alternatively spliced peptide segments. Biochemistry. 1989 Apr 18;28(8):3293–3298. doi: 10.1021/bi00434a026. [DOI] [PubMed] [Google Scholar]
  41. Tamkun J. W., Hynes R. O. Plasma fibronectin is synthesized and secreted by hepatocytes. J Biol Chem. 1983 Apr 10;258(7):4641–4647. [PubMed] [Google Scholar]
  42. Tamkun J. W., Schwarzbauer J. E., Hynes R. O. A single rat fibronectin gene generates three different mRNAs by alternative splicing of a complex exon. Proc Natl Acad Sci U S A. 1984 Aug;81(16):5140–5144. doi: 10.1073/pnas.81.16.5140. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Vibe-Pedersen K., Magnusson S., Baralle F. E. Donor and acceptor splice signals within an exon of the human fibronectin gene: a new type of differential splicing. FEBS Lett. 1986 Oct 27;207(2):287–291. doi: 10.1016/0014-5793(86)81506-x. [DOI] [PubMed] [Google Scholar]
  44. Wayner E. A., Garcia-Pardo A., Humphries M. J., McDonald J. A., Carter W. G. Identification and characterization of the T lymphocyte adhesion receptor for an alternative cell attachment domain (CS-1) in plasma fibronectin. J Cell Biol. 1989 Sep;109(3):1321–1330. doi: 10.1083/jcb.109.3.1321. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES