Internalization but not binding of thrombospondin-1 to low density lipoprotein receptor-related protein-1 requires heparan sulfate proteoglycans.
The amino-terminal domain of the extracellular matrix (ECM) protein thrombospondin-1 (TSP-1) mediates binding to cell surface heparan sulfate proteoglycans (HSPG) as well as binding to the endocytic receptor, low density lipoprotein-related protein (LRP-1). We previously found that recombinant TSP-1 containing the amino-terminal residues 1-214, retained both of these interactions (Mikhailenko et al. [1997]: J Biol Chem 272:6784-6791). Here, we examined the activity of a recombinant protein containing amino-terminal residues 1-90 of TSP-1 and found that this domain did not retain high-affinity heparin-binding. The loss of heparin-binding correlated with decreased binding to the fibroblast cell surface. However, both ligand blotting and solid phase binding studies indicate that this truncated fragment of TSP-1 retained high-affinity binding to LRP-1. Consistent with this, it also retained the ability to block the uptake and degradation of (125)I-TSP-1. However, TSP-1(1-90) itself was poorly endocytosed and this truncated amino-terminal domain was considerably more effective than the full-length heparin-binding domain (HBD) of TSP-1 in blocking the catabolism of endogenously expressed TSP-1. These results indicate that TSP-1 binding to LRP-1 does not require prior or concomitant interaction with cell surface HSPG but suggest subsequent endocytosis requires high-affinity heparin-binding.
Wang
Shuxia
S
Department of Vascular Biology, Jerome H. Holland Laboratory, American Red Cross, Rockville, Maryland 20855, USA. swang@path.uab.edu
Herndon
Mary E
ME
Ranganathan
Sripriya
S
Godyna
Svetlana
S
Lawler
Jack
J
Argraves
W Scott
WS
Liau
Gene
G
D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D019812:Heparan Sulfate Proteoglycans / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006493:Heparin / Q000378:metabolism; D006801:Humans; D026503:Low Density Lipoprotein Receptor-Related Protein-1 / Q000378:metabolism*; D051379:Mice; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D011994:Recombinant Proteins / Q000235:genetics; D019700:Thrombospondin 1 / Q000737:chemistry* / Q000378:metabolism* / Q000494:pharmacology
D017077:Culture Media, Conditioned; D019812:Heparan Sulfate Proteoglycans; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D019700:Thrombospondin 1; D006493:Heparin