Role of A-kinase anchoring proteins (AKAPs) in activation of the sickle cell disease red blood cell adhesion receptor BCAM/Lu
Jamie Maciaszek (University of Connecticut), Biree Andemariam (University of Connecticut Health Center), George Lykotrafitis (University of Connecticut)
Experimental Nanobiomechanics
Mon 9:00 - 10:30
Barus-Holley 163
The abnormal adherence of sickle RBCs (SS RBCs) to the vascular endothelium is a major contributor to painful vaso-occlusive episodes. Several SS RBC surface receptors interact with the endothelial wall components to mediate vascular adhesion, with one important interaction being between BCAM/Lu on the RBC with the 5 chain of laminin (laminin-5), a component of the extracellular matrix of the vascular endothelium. Here, we investigated whether A-kinase anchoring proteins (AKAPs) play a role in the downstream regulation of BCAM/Lu adhesion to laminin-5 through the PKA pathway. Single-cell atomic force microscopy (AFM) was employed to identify the frequency and binding force of the RBC adhesion receptor BCAM/Lu with laminin-5. Molecular interactions were quantified by recording force versus distance curves between a laminin-5 functionalized probe and the RBC surface. Binding forces were plotted as a dot map using MATLAB to display the discrete location of BCAM/Lu receptors on the RBC surface, and were also plotted as a frequency distribution and fit with the Gaussian model to obtain the average magnitude of the binding force between BCAM/Lu and laminin-5. To determine if AKAPs play a role in the downstream regulation of BCAM/Lu adhesion to laminin-5 through the PKA pathway, we used a St-Ht31 inhibitor peptide. We found that SS RBC adhesion to laminin-5 was reduced to various extents in 3 patients (0.40, 0.13, and 0.61 fold). The frequency of BCAM/Lu binding events detected on SS RBCs decreased significantly from 6.88 ± 2.65% at baseline to 1.91 ± 0.28% following St-Ht31 inhibition (p < 0.01). The binding force between BCAM/Lu on the SS RBC and laminin-5 on the AFM probe was unchanged, measuring 43.88 ± 8.87 pN at baseline versus 41.31 ± 10.89 pN following inhibition with AKAP inhibitor St-Ht31. This finding provides evidence for a new paradigm in AKAP adhesive signaling, as well as elucidating a novel signaling pathway that promotes SS RBC adhesion.