Expression and function of the collagen receptor GPVI during megakaryocyte maturation.
In this report, the expression and function of the platelet collagen receptor glycoprotein VI (GPVI) were studied in human megakaryocytes during differentiation and maturation of mobilized blood and cord blood derived CD34(+) cells. By flow cytometry, using an anti-GPVI monoclonal antibody or convulxin, a GPVI-specific ligand, GPVI was detected only on CD41(+) cells including some CD41(+)/CD34(+) cells, suggesting expression at a stage of differentiation similar to CD41. These results were confirmed at the mRNA level using reverse transcription-polymerase chain reaction. GPVI expression was low during megakaryocytic differentiation but increased in the more mature megakaryocytes (CD41(high)). As in platelets, megakaryocyte GPVI associates with the Fc receptor gamma chain (FcRgamma). The FcR gamma chain was detected at the RNA and protein level at all stages of megakaryocyte maturation preceding the expression of GPVI. The other collagen receptor, alpha(2)beta(1) integrin (CD49b/CD29), had a pattern of expression similar to GPVI. Megakaryocytic GPVI was recognized as a 55-kDa protein by immunoblotting and ligand blotting, and thus it presented a slightly lower apparent molecular mass than platelet GPVI (58 kDa). Megakaryocytes began to adhere to immobilized convulxin via GPVI after only 8-10 days of culture, at a time when megakaryocytes were maturing. At this stage of maturation, they also adhered to immobilized collagen by alpha(2)beta(1) integrin-dependent and -independent mechanisms. Convulxin induced a very similar pattern of protein tyrosine phosphorylation in megakaryocytes and platelets including Syk, FcRgamma, and PLC(gamma)2. Our results showed that GPVI is expressed early during megakaryocytic differentiation but functionally allows megakaryocyte adherence to collagen only at late stages of differentiation when its expression increases.
INSERM E9907, Faculté Xavier Bichat, 75870 Paris Cedex 18, Paris, France.
A H
AH
Lagrue-Lak-Hal
N
N
Debili
G
G
Kingbury
C
C
Lecut
J P
JP
Le Couedic
J L
JL
Villeval
M
M
Jandrot-Perrus
W
W
Vainchenker
D018952:Antigens, CD34 / Q000378:metabolism; D001483:Base Sequence; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism*; D017931:DNA Primers; D005434:Flow Cytometry; D016023:Integrins / Q000378:metabolism; D008533:Megakaryocytes / Q000276:immunology / Q000378:metabolism*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism; D010980:Platelet Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D038981:Receptors, Collagen; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018952:Antigens, CD34; D017931:DNA Primers; D016023:Integrins; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D010980:Platelet Membrane Glycoproteins; D012333:RNA, Messenger; D038981:Receptors, Collagen; C098341:platelet membrane glycoprotein VI; D003094:Collagen