[Effect of glycoprotein alpha II bA2334C mutation on the biosynthesis and transportation of alpha II bbeta3 complex].
OBJECTIVE
To study the effect of glycoprotein (GP) alpha II bA2334C mutation on the biosynthesis and expression of alpha II bbeta3 complex.


METHODS
The GP alpha II bA2334C eukaryotic expression plasmid pc3.1-2334M2b was constructed. Chinese hamster ovary (CHO) cells were transfected with the plasmid with or without integrin beta3 expression plasmid pc3.1-3a. The whole expression of alpha II bA2334C was confirmed by Western blot and the membrane expression was analyzed by flow cytometry. A newly constructed alpha II bA2334C GFP fusion protein expressing plasmid was used to determine its subcellular localization by laser confocal scanning microscopy.


RESULTS
Expression of the mutant protein, alpha II bA2334C, in the transfected CHO cells was confirmed by Western blot with a lower rate of the mature type than the wild type control. The expression on membrane was only 25% of the normal. Subcellular localization analysis showed that alpha II bA2334C GFP was able to be expressed in CHO cells and could be transported from endoplasmic reticulum to Golgi apparatus.


CONCLUSIONS
The mutant alpha II bA2334C can be synthesized in CHO cells and form alpha II bbeta3 complex. However, only a small fraction of the premature alpha II bA2334C can be transported to Golgi apparatus and transformed to mature alpha II b. The possible pathogenesis of this type II thrombasthenia may be that the misfolded alpha II bA2334C is partially degraded in the endoplasmic reticulum causing lower expression of alpha II bbeta3 complex on the membrane and resulting in impared function of platelets than normal alpha II b.
Fu
Bin
B
Department of Hematology, Xiangya Hospital, National Laboratory of Medical Genetics, Central South University, Changsha 410008, China.
Chen
Fang-ping
FP
Xia
Kun
K
Fu
Gan
G
Liu
Wei
W
Huang
Xi-lian
XL
Xiao
Guang-fen
GF
D000818:Animals; D001692:Biological Transport; D015153:Blotting, Western; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008081:Liposomes; D018613:Microscopy, Confocal; D008875:Middle Aged; D009154:Mutation; D010957:Plasmids / Q000235:genetics; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000235:genetics* / Q000378:metabolism*; D014162:Transfection
D008081:Liposomes; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D049452:Green Fluorescent Proteins