Association of cystic fibrosis transmembrane conductance regulator and protein phosphatase 2C

Cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel s are rapidly deactivated by a membrane-bound phosphatase activity. The eff iciency of this regulation suggests CFTR and protein phosphatases may be as sociated within a regulatory complex. In this paper we test that possibilit y using coimmunoprecipitation and cross-linking experiments. A monoclonal a nti-CFTR antibody co-precipitated type 2C protein phosphatase (PP2C) from b aby hamster kidney cells stably expressing CFTR but did not co-precipitate PP1, PP2A, or PP2B. Conversely, a polyclonal anti-PP2C antibody co-precipit ated CFTR from baby hamster kidney membrane extracts. Exposing baby hamster kidney cell lysates to dithiobis (sulfosuccinimidyl propionate) caused the cross-linking of histidine-tagged CFTR (CFTRHis10) and PP2C into high mole cular weight complexes that were isolated by chromatography on Ni2+-nitrilo triacetic acid-agarose. Chemical cross-linking was specific for PP2C, becau se PP1, PP2A, and PP2B did not co-purify with CFTRhis10 after dithiobis (su lfosuccinimidyl propionate) exposure. These results suggest CFTR and PP2C e xist, in a stable complex that facilitates regulation of the channel.