Cloning and characterization of the annexin II receptor on human marrow stromal cells

Ganwei Lu, Hidefumi Maeda, Sakamuri V. Reddy, Noriyoshi Kurihara, Robin Leach, Judith L. Anderson, G. David Roodman

Research output: Contribution to journalArticle

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Abstract

Annexin II is a heterotetramer, consisting of two 11-kDa (p11) and two 36-kDa (p36) subunits, that is produced by osteoclasts and stimulates osteoclast formation. However, its receptor is unknown. We showed that annexin II binds to normal primary human marrow stromal cells and the Paget's marrow-derived PSV10 stromal cell line to induce osteoclast formation. 125I-Labeled annexin II binding assays with PSV10 cells demonstrated that there was a single class of annexin II receptors with a Kd of 5.79 nM and B max of 2.13 × 105 receptors/cell. Annexin III or annexin V did not bind this receptor. Using 125I-labeled annexin II binding to screen NIH3T3 transfected with a human marrow cDNA expression library, we identified a putative annexin II receptor clone, which encoded a novel 26-kDa type I membrane receptor protein when expressed in HEK 293 cells. HEK 293 cells transformed with the cloned annexin II receptorcDNAshowed a similar binding affinity to annexin II as that observed in PSV10 cells. Chemical cross-linking experiments with biotinylated annexin II and intact PSV10 cells identified a 55-kDa band on Western blot analysis that reacted with both an anti-p11 antibody and streptavidin but not anti-p36 antibody. A rabbit polyclonal antibody raised against the putative recombinant annexin II receptor also recognized the same 26-kDa protein band detected in PSV10 cells. Importantly, the annexin II receptor antibody dose-dependently blocked the stimulatory effects of annexin II on human osteoclast formation, demonstrating that the receptor mediates the effects of annexin II on osteoclast formation.

Original languageEnglish (US)
Pages (from-to)30542-30550
Number of pages9
JournalJournal of Biological Chemistry
Volume281
Issue number41
DOIs
StatePublished - Oct 13 2006
Externally publishedYes

Fingerprint

Annexin A2
Cloning
Stromal Cells
Organism Cloning
Bone Marrow
Osteoclasts
Antibodies
HEK293 Cells
human annexin II receptor
Annexin A3
Anti-Idiotypic Antibodies
Streptavidin
Annexin A5
Gene Library

ASJC Scopus subject areas

  • Biochemistry

Cite this

Cloning and characterization of the annexin II receptor on human marrow stromal cells. / Lu, Ganwei; Maeda, Hidefumi; Reddy, Sakamuri V.; Kurihara, Noriyoshi; Leach, Robin; Anderson, Judith L.; Roodman, G. David.

In: Journal of Biological Chemistry, Vol. 281, No. 41, 13.10.2006, p. 30542-30550.

Research output: Contribution to journalArticle

Lu, Ganwei ; Maeda, Hidefumi ; Reddy, Sakamuri V. ; Kurihara, Noriyoshi ; Leach, Robin ; Anderson, Judith L. ; Roodman, G. David. / Cloning and characterization of the annexin II receptor on human marrow stromal cells. In: Journal of Biological Chemistry. 2006 ; Vol. 281, No. 41. pp. 30542-30550.
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