Mapping of Network-forming, Heparin-binding, and 
1
1 Integrin-recognition Sites within the 
-Chain Short Arm of Laminin-1 (*)
- Holly Colognato-Pyke(1),
- Julian J. O'Rear(2),
- Yoshihiko Yamada(3),
- Salvatore Carbonetto(4),
- Yi-Shan Cheng(1) and
- Peter D. Yurchenco(1)(§)
- From the (1) Departments of Pathology and
- (2) Microbiology and Molecular Genetics, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854,
- (3) NIDR, National Institutes of Health, Bethesda, Maryland 20892, and
- (4) Center for Neuroscience Research, McGill University, Montreal, Quebec H3A 2B2, Canada
- § To whom correspondence should be addressed: Dept. of Pathology, UMDNJ-Robert Wood Johnson Medical School, 675 Hoes Ln., Piscataway, NJ 08854. Tel.: 908-235-4674; Fax: 908-235-4825.
Abstract
Cell-interactive and architecture-forming functions are associated with the short arms of basement membrane laminin-1. To
map and characterize these functions, we expressed recombinant mouse laminin-1 α-chain extending from the N terminus through
one third of domain IIIb. This dumbbell-shaped glycoprotein (rα1(VI-IVb)′), secreted by mammalian cells, was found to possess
three activities. 1) Laminin polymerization was quantitatively inhibited by recombinant protein, supporting an α-chain role
for a three-short arm interaction model of laminin self-assembly. 2) rα1(VI-IVb)′ bound to heparin, and the activity was localized
to a subfragment corresponding to domain VI by 
I-heparin blotting. 3) PC12 rat pheochromocytoma cells adhered to, and rapidly extended branching neurites on, rα1(VI-IVb)′,
with adhesion inhibited by α1 and β1 integrin chain-specific antibodies. The ability of anti-laminin antibody to block PC12
cell adhesion to laminin was selectively prevented by absorption with rα1(VI-IVb)′ or α-chain domain VI fragment. This active
integrin-recognition site could furthermore be distinguished from a second cryptic α1β1-binding site exposed by heat treatment
of fragment P1′, a short arm fragment lacking globules. Thus, a polymer-forming, a heparin-binding, and the active α1β1 integrin-recognition
site are all clustered at the end of the α-chain short arm, the latter two resident solely in domain VI.
Footnotes
-
↵* This study was supported in part by National Institutes of Health Grant R01-DK36425. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
-
↵1 The abbreviations used are:
- EHS
-
Engelbreth-Holm-Swarm
- DMEM
-
Dulbecco's modified Eagle's medium
- PMSF
-
phenylmethylsulfonyl fluoride
- HPLC
-
high performance liquid chromatography
- PAGE
-
polyacrylamide gel electrophoresis
- ELISA
-
enzyme-linked immunosorbent assay
- TBS
-
Tris-buffered saline
- D-PBS
-
Dulbecco's phosphate-buffered saline
- NGF
-
nerve growth factor
- kb
-
kilobase(s).
- © 1995 by The American Society for Biochemistry and Molecular Biology, Inc.
