A functional link between the chloride channel cystic fibrosis transmembrane conductance regulator (CFTR) and the epithelial Na+ channel (ENaC), has been suggested based on evidence from electrophysiological and biochemical methods in heterologous expression systems. Their interaction in native tissue is not clearly defined but is implicated in the pathophysiology of cystic fibrosis. The present study utilized immunofluorescence and laser confocal microscopy on cryosections of healthy human skin specimens to characterize CFTR and ENaC localization in sweat ducts. Ductal segment morphology was identified with 4',6‐diamidino‐2‐phenylindole nuclear staining. Throughout the duct there was one to two fold less staining of CFTR compared to ENaC. The proportion of lumen‐specific staining was significantly (one to ten fold) greater for CFTR than for the more ubiquitous ENaC, but varied widely between tissue samples. There is additional corroborating evidence of some co‐localization of CFTR and ENaC confined primarily to the ductal lumen, a principal site of transepithelial NaCl reabsorption. This study is the first example of immunofluorescent staining for CFTR and ENaC localization in native human tissue. Future studies will help identify the spatial relationship between these two channel proteins, and how their potential interaction correlates to fluid and electrolyte transport in health and disease.
G Protein‐coupled receptors (GPCRs) are the third largest family of genes in the human genome and are the target of many pharmaceuticals. Evidence has shown that many GPCRs form heteromeric complexes, although the functional consequences of such interactions are unclear. Bradykinin type 2 receptor (Bk2R) and Beta 2 adrenergic receptor ( β2AR), Gαq‐ and Gαs‐coupled receptors, respectively, are GPCRs that can heterodimerize with other members of their classes in vitro and in vivo, participate in cardiovascular regulation, and are coexpressed in many cell types, including pheochromocytoma (PC12) cells. Given these factors, we examined potential interplay between Bk2R and β2AR. We hypothesized that Bk2R and β2AR physically associate, changing native receptor signaling. Co‐immunoprecipitation studies in PC12 cells and X. laevis oocytes showed physical association of Bk2R and β2AR. In both PC12 cells and oocytes expressing these receptors, bradykinin‐induced Bk2R transactivation via both Gαs‐ and Gαq‐coupled pathways, which was significantly ablated by inverse agonists of Bk2R or β2AR; thus, conformational changes in both receptors are required for transactivation. This is the first evidence of Bk2R/β2AR physical interaction and functional cross‐talk. Given the relevance of pharmaceutics that affect Bk2R and β2AR, these exciting data may provide opportunities for drug development and refinement.
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