Dependence on vitamin K-dependent protein S for eukaryotic cell secretion of the beta-chain of C4b-binding protein.
The anticoagulant vitamin K-dependent protein S (PS) circulates in plasma in two forms, 30% free and 70% being bound to the complement regulatory protein C4b-binding protein (C4BP). The major C4BP isoform consists of 7 α-chains and 1 β-chain (C4BPβ(+)), the chains being linked by disulfide bridges. PS binds to the β-chain with high affinity. In plasma, PS is in molar excess over C4BPβ(+) and due to the high affinity, all C4BPβ(+) molecules contain a bound PS. Taken together with the observation that PS-deficient patients have decreased levels of C4BPβ(+), this raises the question of whether PS is important for secretion of the β-chain from the cell. To test this hypothesis, HEK293 cells were stably and transiently transfected with β-chain cDNA in combinations with cDNAs for PS and/or the α-chain. The concentration of β-chains in the medium increased after co-transfection with PS cDNA, but not by α-chain cDNA, suggesting secretion of the β-chains from the cells to be dependent on concomitant synthesis of PS, but not of the α-chains. Thus, β-chains that were not disulfide-linked to the α-chains were secreted in complex with PS, either as monomers or dimers. Pulse-chase demonstrated that the complexes between PS and β-chain were formed intracellularly, in the endoplasmic reticulum. In conclusion, our results demonstrate that successful secretion of β-chains depends on intracellular complex formation with PS, but not on the α-chains. This provides an explanation for the decreased β-chain levels observed in PS-deficient patients.
Carlsson
Sofia
S
Department of Laboratory Medicine, Division of Clinical Chemistry, Lund University, Skåne University Hospital, SE-205 02 Malmö, Sweden.
Dahlbäck
Björn
B
D000818:Animals; D001464:Barium / Q000378:metabolism; D002460:Cell Line; D050716:Complement C4b-Binding Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005057:Eukaryotic Cells / Q000378:metabolism*; D006801:Humans; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017293:Protein S / Q000235:genetics / Q000378:metabolism*; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D050716:Complement C4b-Binding Protein; D020033:Protein Isoforms; D017293:Protein S; D021122:Protein Subunits; D001464:Barium