(596u) Developing a Quantitative Method to Investigate Receptor Oligomerization: Applications to Receptor for Advanced Glycation Endproducts (RAGE)

Transmembrane (TM) receptors play a key role in cell-cell communication and signal transduction that regulates diverse biological processes including cell migration, cell proliferation, and cellular differentiation. In most cases, regulation of receptor signaling occurs through receptor oligomerization, in which the interplay between interactions involving multiple receptor domains are necessary for stabilizing the activated form of the receptor for transmembrane signaling to occur. Thus, there is strong correlation observed between specific interactions involving transmembrane and cytoplasmic domains and receptor activation.  While there are several tools available to investigate receptor oligomerization in mammalian (BRET) and bacterial (TOXCAT) membranes, technical challenges associated with these assays can limit their utility in rapidly analyzing regions of receptors responsible for oligomerization. Therefore, we developed a robust, transcriptional activator-based method to measure receptor domain oligomerization in bacterial membranes, and benchmarked this method against integrin α_IIb TM-CYTO with known mutations that promotes homooligomerization. We then extended this method to identify regions of the receptor for advanced glycation endproducts (RAGE) that promote ligand-independent oligomerization. RAGE receptor has been associated with several chronic disorders such as Alzheimer’s disease, atherosclerosis, diabetes, and cancers. Furthermore, several studies have shown that RAGE homodimerization is essential for ligand recognition and signal transduction. Our results point to a key role for the RAGE cytosolic domain in promoting ligand-independent oligomerization, in contrast to previous studies that focused primarily on the soluble, secreted ectodomain.