LAUSR

This report identifies a novel variant form of the inherited bleeding disorder, Glanzmann thrombasthenia (GT) exhibiting only mild bleeding in a physically active individual. The platelets cannot aggregate ex vivo with physiologic agonists of activation, although microfluidic analysis with whole blood displays moderate ex vivo platelet adhesion and aggregation consistent with mild bleeding. Immunocytometry shows reduced expression of αIIbβ3 on quiescent platelets that spontaneously bind/store fibrinogen and activation-dependent antibodies (LIBS-319.4, PAC-1) report β3 extension suggesting an intrinsic activation phenotype. Genetic analysis reveals a single F153Sβ3 substitution within the βI-domain from a heterozygous T556C nucleotide substitution of ITGB3 exon 4 in conjunction with a previously reported IVS5(+1)G>A splice-site mutation with undetectable platelet mRNA accounting for hemizygous expression of F153Sβ3. F153 is completely conserved among β3 of several species and all human β-integrin subunits suggesting that it may play a vital role in integrin structure/function. Mutagenesis of αIIb-F153β3 also displays reduced-levels of a constitutively activated αIIb-S153β3 on HEK293T cells. The overall structural analysis suggests that a bulky-aromatic, non-polar amino acid (AA) (F,W)153β3 is critical for maintaining the resting conformation of α2- and α1-helixes of the βI-domain because small AA substitutions (S,A) facilitate an unhindered inward movement of the α2- and α1-helixes of the βI-domain towards the constitutively active αIIbβ3 conformation, while a bulky-aromatic, polar AA (Y) hinders such movements and restrains αIIbβ3 activation. The data collectively demonstrate that disruption of F153β3 can significantly alter normal integrin/platelet function, although reduced expression of αIIb-S153β3 maybe compensated by a hyperactive conformation that promotes viable hemostasis.