Elucidating the role of junctional adhesion molecule‐A homodimerization in angiogenesis

Angiogenesis is the formation of new blood vessels from pre‐existing ones. It is involved in both physiological and pathological processes. Junctional adhesion molecule‐A (JAM‐A) is required for the induction of angiogenesis by basic fibroblast growth factor. JAM‐A is also known to form homodimers in vitro and in vivo . This study intends to determine the relationship between JAM‐A homodimerization and angiogenesis. For this purpose, site‐directed mutagenesis was performed on JAM‐A cDNA constructs to introduce mutations designed to impair homodimerization. Our goal is to use these constructs to examine the relationship between angiogenesis and JAM‐A homodimerization in a blood vessel endothelial cell model. In order to confirm that the mutations do, in fact, impair homodimerization, Chinese hamster ovary (CHO) cells were transfected with wild‐type or mutant JAM‐A constructs. The cells were then treated with a cross‐linker, bis(sulfosuccinimidyl) suberate (BS 3 ), lysed, and subjected to Western blotting for JAM‐A. Putative homodimer bands were seen at roughly twice the molecular weight of monomeric JAM‐A. Dimer bands were detected at similar intensities for both mutant and wild‐type JAM‐A, indicating no difference in homodimerization. A different approach may be needed to detect any differences. This research was funded by the Arnold and Mabel Beckman Foundation and the Barry M. Goldwater Foundation.