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

Vinata B. Lokeshwar, Nevis L. Fregien, Lilly Y W Bourguignon

Research output: Contribution to journalArticle

187 Citations (Scopus)

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 (NGGNGTVEDRKPSEL) containing 15 aa between aa 305 and aa 320, support the conclusion that this region is required for ankyrin 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 ankyrin- 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 (≥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.

Original languageEnglish
Pages (from-to)1099-1109
Number of pages11
JournalJournal of Cell Biology
Volume126
Issue number4
DOIs
StatePublished - Aug 1 1994

Fingerprint

Ankyrins
Hyaluronic Acid
Proteins
COS Cells
Cell Adhesion Molecules
Cell Adhesion
Extracellular Matrix
Protein Isoforms
Complementary DNA
Binding Sites
Amino Acids

ASJC Scopus subject areas

  • Cell Biology

Cite this

Ankyrin-binding domain of CD44(GP85) is required for the expression of hyaluronic acid-mediated adhesion function. / Lokeshwar, Vinata B.; Fregien, Nevis L.; Bourguignon, Lilly Y W.

In: Journal of Cell Biology, Vol. 126, No. 4, 01.08.1994, p. 1099-1109.

Research output: Contribution to journalArticle

@article{beb035dbefdd47688bfb171a54df441d,
title = "Ankyrin-binding domain of CD44(GP85) is required for the expression of hyaluronic acid-mediated adhesion function",
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 (NGGNGTVEDRKPSEL) containing 15 aa between aa 305 and aa 320, support the conclusion that this region is required for ankyrin 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 ankyrin- 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 (≥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.",
author = "Lokeshwar, {Vinata B.} and Fregien, {Nevis L.} and Bourguignon, {Lilly Y W}",
year = "1994",
month = "8",
day = "1",
doi = "10.1083/jcb.126.4.1099",
language = "English",
volume = "126",
pages = "1099--1109",
journal = "Journal of Cell Biology",
issn = "0021-9525",
publisher = "Rockefeller University Press",
number = "4",

}

TY - JOUR

T1 - Ankyrin-binding domain of CD44(GP85) is required for the expression of hyaluronic acid-mediated adhesion function

AU - Lokeshwar, Vinata B.

AU - Fregien, Nevis L.

AU - Bourguignon, Lilly Y W

PY - 1994/8/1

Y1 - 1994/8/1

N2 - 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 (NGGNGTVEDRKPSEL) containing 15 aa between aa 305 and aa 320, support the conclusion that this region is required for ankyrin 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 ankyrin- 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 (≥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.

AB - 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 (NGGNGTVEDRKPSEL) containing 15 aa between aa 305 and aa 320, support the conclusion that this region is required for ankyrin 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 ankyrin- 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 (≥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.

UR - http://www.scopus.com/inward/record.url?scp=0028141591&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0028141591&partnerID=8YFLogxK

U2 - 10.1083/jcb.126.4.1099

DO - 10.1083/jcb.126.4.1099

M3 - Article

C2 - 7519619

AN - SCOPUS:0028141591

VL - 126

SP - 1099

EP - 1109

JO - Journal of Cell Biology

JF - Journal of Cell Biology

SN - 0021-9525

IS - 4

ER -