GLANZMANN THROMBASTHENIA DUE TO A 2-AMINO-ACID DELETION IN THE 4TH CALCIUM-BINDING DOMAIN OF ALPHA(IIB) - DEMONSTRATION OF THE IMPORTANCE OF CALCIUM-BINDING DOMAINS IN THE CONFORMATION OF ALPHA(IIB)BETA(3)

The integrin alpha(llb)beta(3), a calcium-dependent heterodimer, plays a critical role in platelet aggregation. The alpha(llb) subunit of th e heterodimer contains four highly conserved putative calcium-binding domains in its extracellular portion. During studies of the molecular basis of Glanzmann thrombasthenia in a child of mixed Caucasian backgr ound whose platelets expressed little alpha(llb)beta(3) on their surfa ce, we found the patient heterozygous for a two amino acid deletion in the fourth alpha(llb) calcium-binding domain. When this alpha(llb) mu tant was expressed in COS-1 cells, we found that the deletion did not interfere with the assembly of alpha(llb)beta(3) heterodimers, but al tered their conformation such that they were neither recognized by the heterodimer-specific antibody A2A9 nor able to undergo further intrac ellular processing or transport to the cell surface. These results sug gest that the calcium-binding domains in alpha(llb) play an important role maintaining the overall conformation of alpha(llb)beta 3. To conf irm this suggestion, we deleted each of the four 12 amino acid calcium -binding domains in alpha(llb) by in vitro mutagenesis and expressed t he mutants along with pg in COS-1 cells. Each construct formed a heter odimer with PSI but none of the heterodimers interacted with A2A9 or u nderwent further intracellular processing. These data indicate that th e calcium-binding domains in alpha(llb) are not involved in alpha(llb) beta(3) heterodimer formation, but their presence is required for the intracellular transport of alpha(llb)beta(3) to the cell surface. (C) 1996 by The American Society of Hematology.