Platelets express three different splice variants of ApoER2 that are all involved in signaling

M.T. Pennings, R.H.W.M. Derksen, R.T Urbanus, W.L.H. Dalessi - Tekelenburg, W. Hemrika, Ph.G. de Groot

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Background: ß2-Glycoprotein I is themost relevant antigen in antiphospholipid syndrome. We have shown that binding of dimerized ß2-GPI to platelets viaApoER2¢ sensitizes platelets for second activating stimuli. Objective: Determine the region of ApoER2 involved in the binding of dimeric b2-GPI. Methods: Cultured human megakaryocytes (MK) and three different human megakaryocytic cell lines were used for mRNA isolation to clone and express recombinant soluble platelet ApoER2. Domain deletion mutants of ApoER2 were constructed to identify the binding site for dimeric ß2-GPI. The presence of ApoER2 splice variants in platelets was demonstrated by immuno-blotting. Results: Three different mRNA splice variants were isolated from all four types of megakaryocytic cells used. Sequence analysis identified the splice variants: (i) shApoER2D5 lacking low-density lipoprotein (LDL) binding domains 4, 5 and 6; (ii) shApoER2D4-5 lacking LDL binding domains 3, 4, 5, 6 and (iii) shApoER2D3-4-5 lacking LDL binding domains 3, 4, 5, 6 and 7. The presence of three splice variants of ApoER2 on platelets was confirmed by immuno-blotting, with ApoER2D4-5 being the most abundantly expressed splice variant. Upon stimulation with dimeric ß2-GPI, all three splice variantswere translocated to the cytosol; however, ApoER2D4-5 translocation was most prominent. Dimeric b2-GPI binds platelet ApoER2 variants via LDLbinding domain 1. Conclusions: Three different ApoER2 mRNA splice variants were isolated from MK and platelets express all three splice variants. All splice variants were shown to be functional by translocation upon stimulationwith dimeric ß2-GPI. All three splice variants express LDL-binding domain 1.
Original languageUndefined/Unknown
Pages (from-to)1538-1544
Number of pages7
JournalJournal of Thrombosis and Haemostasis
Volume5
Issue number7
Publication statusPublished - 2007

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