Abstract

This research represents a continuation of our studies on alpha-D-galactosyl-containing glycoproteins present on certain animal cells and tissues. We will focus attention on the carbohydrate moieties of two basement membrane and cell surface glycoproteins: laminin and entactin, both of which contain alpha-D-galactosyl end groups. Laminin is believed to be present on the surface of highly metastatic cells. Structural and biosynthetic studies will be conducted on the carbohydrate residues of these glycoproteins as well as on the alpha-D-galactosyl-terminated glycoproteins present on the surface of Ehrlich tumor cells. Purified lectins of known carbohydrate binding specificity, antibodies raised against murine laminin and against a trisaccharide hapten present as part of the structure of these glycoproteins, will be employed in these studies. An attempt will be made to protect mice against a lethal dose of Ehrlich cells by immunizing the mice with the trisaccharide-protein conjugate. The appearance of cell surface laminin, which is expressed on stimulated but not resident murine macrophages, will be quantified under various conditions of stimulation using a lectin-enzyme conjugate. Biosynthetic studies will include an investigation of the incorporation of D-galactose and N-acetyl-D-glucosamine into the polylactosamineglycan that forms part of the structure of these alpha-D-galactosyl-terminated glycoproteins. We shall characterize an alpha-D-galactosidase present in Ehrlich ascites tumor cells and that probably is involved in the biodegradation of alpha-D-galactosyl-containing glycoproteins. The localization of alpha-D-galactosyl-terminated glycoconjugates in a variety of cells by electron microscopy will be conducted using a colloidal gold complex of Griffonia simplicifolia I-B?4? isolectin, a probe specific for alpha-D-galactosyl end groups. We shall also attempt to select and characterize a variant of Ehrlich ascites tumor cells that lack alpha-D-galactosyl groups. (A)

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA020424-12
Application #
3165294
Study Section
Physiological Chemistry Study Section (PC)
Project Start
1984-12-01
Project End
1989-11-30
Budget Start
1987-12-01
Budget End
1988-11-30
Support Year
12
Fiscal Year
1988
Total Cost
Indirect Cost

  Institution

Name
University of Michigan Ann Arbor
Department
Type
Schools of Medicine
DUNS #
791277940
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109

  Publications

Roth, J; Goldstein, I J (1995) Subcellular distribution of terminal alpha-D- and beta-D-galactosyl residues in Ehrlich tumour cells studied by lectin-gold techniques. Glycoconj J 12:142-9
Song, Z; Varani, J; Goldstein, I J (1995) Expression and function of beta 1 integrins on adherent and nonadherent Ehrlich ascites tumor cells. Exp Cell Res 218:96-104
Roth, J; Li, W P; Knibbs, R N et al. (1994) Differential expression of cell surface sialoglycoconjugates on wild-type and cultured Ehrlich tumor cells as revealed by quantitative lectin-gold ultrastructural cytochemistry. Proc Natl Acad Sci U S A 91:11353-7
Chen, Y F; Boland, C R; Kraus, E R et al. (1994) The lectin Griffonia simplicifolia I-A4 (GS I-A4) specifically recognizes terminal alpha-linked N-acetylgalactosaminyl groups and is cytotoxic to the human colon cancer cell lines LS174t and SW1116. Int J Cancer 57:561-7
Knibbs, R N; MacCallum, D K; Lillie, J H et al. (1994) Wild-type and cultured Ehrlich ascites tumour cells differ in tumorigenicity, lectin binding patterns and binding to basement membranes. Glycobiology 4:419-28
Shigeta, S; Winter, H C; Goldstein, I J (1994) Alpha-(2-->3)- and alpha-(2-->6)-sialyltransferase activities present in three variants of Ehrlich tumor cells: identification of the products derived from N-acetyllactosamine and beta-D-Gal-(1-->3)-alpha-D-GalNAc-(1-->O)-Bn. Carbohydr Res 264:111-21
Knibbs, R N; Agrwal, N; Wang, J L et al. (1993) Carbohydrate-binding protein 35. II. Analysis of the interaction of the recombinant polypeptide with saccharides. J Biol Chem 268:14940-7
Song, Z; Varani, J; Goldstein, I J (1993) Differences in cell surface carbohydrates, and in laminin and fibronectin synthesis, between adherent and non-adherent Ehrlich ascites tumor cells. Int J Cancer 55:1029-35
Takagaki, M; Knibbs, R N; Roth, J et al. (1993) Monoclonal antibodies that recognize the trisaccharide epitope Gal alpha 1-3Gal beta 1-4GlcNAc present on Ehrlich tumor cell membrane glycoproteins. Histochemistry 100:139-47
Knibbs, R N; Osborne, S E; Glick, G D et al. (1993) Binding determinants of the sialic acid-specific lectin from the slug Limax flavus. J Biol Chem 268:18524-31

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