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  • Osteopontin and the C-terminal peptide of thrombospondin-4 compete for CD44 binding and have opposite effects on CD133+ cell colony formation.

Osteopontin and the C-terminal peptide of thrombospondin-4 compete for CD44 binding and have opposite effects on CD133+ cell colony formation.

BMC research notes (2009-10-27)
Gulzhakhan Sadvakassova, Monica C Dobocan, Luis F Congote
ABSTRACT

C21, the C-terminal peptide of thrombospondin-4, has growth promoting activity and was discovered as one of several erythropoietin-dependent endothelial proteins. C21 stimulates red cell formation in anemic mice and is a growth factor for CD34+ and CD36+ hematopoietic cells, skin fibroblasts and kidney epithelial cells. ROD1 has been identified as an intracellular mediator. Nothing is known about the existence of putative C21 receptors on plasma membranes of target cells. We analyzed the nature of C21-binding proteins in cell lysates of skin fibroblasts using C21 affinity columns. The membrane receptor CD44 was identified as C21-binding protein by mass spectrometry. We were unable to demonstrate any direct involvement of CD44 on cell growth or the effect of C21 on cell proliferation. A soluble form of CD44 was synthesized in insect cells and purified from culture supernatants with a combination of PVDF filtration in the presence of ammonium sulphate and HPLC. Both osteopontin and hyaluronic acid competitively displaced Biotin-C21 binding to CD44. In a colony-forming assay using primitive CD133+ hematopoietic stem cells from cord blood, osteopontin and C21 had opposite effects and C21 reduced the inhibitory action of osteopontin. CD44 is a C21-binding membrane protein. We could not demonstrate an involvement of CD44 in the proliferative action of C21. Nevertheless, based on the antagonism of C21 and osteopontin in hematopoietic precursors, we speculate that C21 could indirectly have a major impact on hematopoietic stem cell proliferation, by hindering osteopontin membrane binding at the level of the bone marrow niche.

MATERIALS
Product Number
Brand
Product Description

SAFC
EX-CELL® 405 Serum-Free Medium for Insect Cells, Protein-free, with L-glutamine, without sodium bicarbonate, dry powder, suitable for insect cell culture
SAFC
EX-CELL® 405 Serum-Free Medium for Insect Cells, with L-glutamine, liquid, sterile-filtered, suitable for insect cell culture