RRC ID 51998
Author Fitter S, Gronthos S, Ooi SS, Zannettino AC.
Title The Mesenchymal Precursor Cell Marker Antibody STRO-1 Binds to Cell Surface Heat Shock Cognate 70.
Journal Stem Cells
Abstract Since its discovery more than 25 years ago, the STRO-1 antibody has played a fundamental role in defining the hierarchical nature of mesenchymal precursor cells (MPC) and their progeny. STRO-1 antibody binding remains a hallmark of immature pluripotent MPC. Despite the significance of STRO-1 in the MPC field, the identity of the antigen has remained elusive. Using a combination of two-dimensional gel electrophoresis, coupled with Western blotting and Tandem mass spectroscopy, we have identified the STRO-1 antigen as heat shock cognate 70 (HSC70;HSPA8). STRO-1 binds to immune-precipitated HSC70 and siRNA-mediated knock down of HSPA8 reduced STRO-1 binding. STRO-1 surface binding does not correlate with HSC70 expression and sequestration of cholesterol reduces STRO-1 surface binding, suggesting that the plasma membrane lipid composition may be an important determinant in the presentation of HSC70 on the cell surface. HSC70 is present on the surface of STRO-1+ but not STRO-1- cell lines as assessed by cell surface biotinylation and recombinant HSC70 blocks STRO-1 binding to the cell surface. The STRO-1 epitope on HSC70 was mapped to the ATPase domain using a series of deletion mutants in combination with peptide arrays. Deletion of the first four amino acids of the consensus epitope negated STRO-1 binding. Notably, in addition to HSC70, STRO-1 cross-reacts with heat shock protein 70 (HSP70), however all the clonogenic cell activity is restricted to the STRO-1BRIGHT /HSP70- fraction. These results provide important insight into the properties that define multipotent MPC and provide the impetus to explore the role of cell surface HSC70 in MPC biology. Stem Cells 2017;35:940-951.
Volume 35(4)
Pages 940-951
Published 2017-4
DOI 10.1002/stem.2560
PMID 28026090
MeSH Amino Acid Sequence Antibodies / metabolism* Antigens, Surface / metabolism* Biomarkers / metabolism Bone Marrow Cells / cytology Cell Line Cholesterol / metabolism Colony-Forming Units Assay Epitope Mapping HSC70 Heat-Shock Proteins / chemistry HSC70 Heat-Shock Proteins / metabolism* HSP70 Heat-Shock Proteins / metabolism Humans Leukocytes, Mononuclear / cytology Membrane Microdomains / metabolism Mesenchymal Stromal Cells / cytology Mesenchymal Stromal Cells / metabolism* Protein Binding Protein Domains
IF 5.587
Resource
Human and Animal Cells UE7T-13(RCB2161)