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PMID: 7519619 Published · ppublish English Comparative Study Journal Article Research Support, U.S. Gov't, P.H.S.

Ankyrin-binding domain of CD44(GP85) is required for the expression of hyaluronic acid-mediated adhesion function.

The Journal of cell biology ·Vol. 126 ·No. 4 ·1994-08-00 ·Pages 1099-109

Lokeshwar VB, Fregien N, Bourguignon LY

Abstract

GP85 is one of the most common hemopoietic isoforms of the cell adhesion molecule, CD44. CD44(GP85) is known to contain at least one ankyrin-binding site within its 70 aa cytoplasmic domain and to bind hyaluronic acid (HA) with its extracellular domain. In this study we have mapped the ankyrin-binding domain of CD44(GP85) by deleting various portions of the cytoplasmic region followed by expression of these truncated cDNAs in COS cells. The results of these experiments indicate that the ankyrin-binding domain resides between amino acids 305 and 355. Biochemical analyses, using competition binding assays and a synthetic peptide (NGGNGT-VEDRKPSEL) containing 15 aa between aa 305 and aa 320, support the conclusion that this region is required for ankryin binding. Furthermore, we have constructed a fusion protein in which this 15 aa sequence of CD44(GP85) is transplanted onto another transmembrane protein which does not bind ankyrin. Our results show that this fusion protein acquires the ability to bind ankyrin confirming that the sequence (306NGGNGTVEDRKPSE320L) is a critical part of the ankryin-binding domain of CD44(GP85). In addition, we have demonstrated that deletion of this 15 aa ankyrin-binding sequence from CD44(GP85) results in a drastic reduction (> or = 90%) of HA-binding and HA-mediated cell adhesion. These findings strongly suggest that ankyrin binding to the cytoplasmic domain of CD44(GP85) plays a pivotal role in regulating hyaluronic acid-mediated cell-cell and cell-extracellular matrix interactions.

MeSH Terms
Amino Acid Sequence Animals Ankyrins/metabolism Base Sequence Binding Sites Carrier Proteins/biosynthesis,metabolism Cattle Cell Adhesion/drug effects,physiology Cell Line Chlorocebus aethiops Cricetinae Horses Humans Hyaluronan Receptors Hyaluronic Acid/metabolism,pharmacology Kidney Kinetics Leukocyte Common Antigens/biosynthesis Mice Molecular Sequence Data Oligodeoxyribonucleotides Rats Receptors, Cell Surface/biosynthesis,metabolism Receptors, Lymphocyte Homing/biosynthesis,metabolism Recombinant Fusion Proteins/biosynthesis Recombinant Proteins/biosynthesis,metabolism Sequence Deletion Sequence Homology, Amino Acid Transfection
Chemicals
Ankyrins Carrier Proteins Hyaluronan Receptors Oligodeoxyribonucleotides Receptors, Cell Surface Receptors, Lymphocyte Homing Recombinant Fusion Proteins Recombinant Proteins Hyaluronic Acid Leukocyte Common Antigens
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Lokeshwar V B
Department of Cell Biology and Anatomy, University of Miami School of Medicine, Florida 33101.
Fregien N
Bourguignon L Y
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.126.4.1099
Published
1994-08-00
Pages
1099-109
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2120123
Subset
IM
Grants
NCI NIH HHS · 1F32CA06057 · United States
NIGMS NIH HHS · GM 36353 · United States
Databases
GENBANK
X6682
External Links
PubMed source DOI: 10.1083/jcb.126.4.1099 PMC Full Text View journal details
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