Common human alpha2a&c adrenergic receptor polymorphisms do not affect cellular processing nor surface trafficking

Abstract

Naturally occurring variant genetic forms of α2a and/or α2c adrenergic receptors (α2a&c ARs) have been identified and linked to congestive heart failure. Two well described α2a&c AR variants occur within the 3rd intracellular loop, α2aLys251 and α2cDel322‐325. We investigated if these variant forms exhibited alterations in localization or cellular processing to account for their phenotypic differences.Human cDNA encoding wild‐type and polymorphic α2a&c ARs were either purchased commercially or created using standard Quick Change mutagenesis techniques, incorporating an amino terminal HA epitope. Each construct was examined with respect to drug binding, biochemical processing (e.g. glycosylation), and surface and total expression (e.g. immunocytochemistry and FACS analysis).All four cDNA constructs were successfully transfected and expressed in HEK293 cells. RX 821002 binding did not differ greatly between wild‐type and polymorphic forms. Transfected cells were analyzed by immunocytochemistry and FACS; neither polymorphic form demonstrated differences in cell surface or intracellular expression patterns. Lastly, endoglycosidase digestions of transfected cell membranes revealed that the polymorphic forms were processed in a similar manner as their wild‐type counterparts.It has been shown previously that certain GPCR polymorphisms can affect receptor stability and trafficking. We examined the two most common α2a&c AR polymorphisms and found no discernable difference in receptor expression, drug binding, surface trafficking, or biochemical processing. Therefore, phenotypic differences between wild‐type and polymorphic forms are most likely due to signal transduction, or possibly neuronal specific influences.This work was supported in part by NINDS K08 NS050654‐01A1 (TA)