HRG TOKUSHIMA - MOLECULAR AND CELLULAR CHARACTERIZATION OF HISTIDINE-RICH GLYCOPROTEIN (HRG) DEFICIENCY

Previously, we found the first congenital deficiency of histidine-rich glycoprotein (HRG) in a Japanese woman with thrombosis. To elucidate the genetic basis of this deficiency, we first performed Southern blot analysis and found no gross deletion or insertion in the proband's HR G gene, We then examined the nucleotide sequences of all seven exons o f the proband's HRG gene. A single nucleotide substitution, G to A at nucleotide position 429, which mutates Gly85 to Glu in the first cysta tin-like domain, was found in exon 3 in 13 of 22 amplified clones, Thi s mutation generates a unique Tao I site, Exon 3 was amplified from th e proband, her family members, and 50 unrelated normal Japanese indivi duals, and Tao I fragmentation was examined, Fragmentation of exon 3 w as observed in one allele of the genes from the proband and the family members who also have decreased plasma levels of HRG. Fifty unrelated normal Japanese individuals had a normal HRG gene, indicating that th e G to A mutation is not a common polymorphism, To elucidate the ident ified mutation as a cause for the secretion defect of HRG in the proba nd's plasma, we constructed and transiently expressed the recombinant Tokushima-type HRG mutant (Gly85 to Glu) in baby hamster kidney (BHK) cells, and examined an intracellular event of the mutant protein, The results showed that only about 20% of the Tokushima-type HRG was secre ted into the culture medium, and intracellular degradation of the muta nt was observed. Thus, the present study strongly suggests that the HR G deficiency is caused by intracellular degradation of the Gly85 to Gl u mutant of HRG in the proband. (C) 1998 by The American Society of He matology.