ΔF508 CFTR Expressed In Xenopus Oocytes Exhibits Unique Thermal Sensitivity

Abstract

The deletion of a phenylalanine at position 508 is the most common, disease-related mutation in the CFTR protein. ΔF508 CFTR channels are assembled in mammalian cells, but exhibit two deficiencies thought to underlie the disease phenotype: impaired trafficking and defective gating. The CFTR trafficking defect is partially mitigated at low temperature so that Xenopus oocytes are ideal for studying the physical properties of ΔF508 channels under physiological conditions. In order to investigate possible effects of the Phe deletion on the intrinsic stability of CFTR channel function in the plasma membrane, we monitored channel activity at room temperature (∼23°C) and during a brief (10-12 minute) period of elevated temperature (28°C to 37°C). In oocytes expressing wild type CFTR, a temperature challenge resulted in a reversible increase in the conductance; a result of a simultaneous increase in single-channel conductance and open probability. Both parameters, however, returned to baseline values when the temperature was returned to 23°C. In marked contrast, conductance due to ΔF508 CFTR channels exhibited a transient increase within a minute after the temperature challenge, followed by a quasi-exponential decline of about 80-90% of the initial conductance (t1/2 = 4 minutes at 37°C). The temperature induced decrease in ΔF508 CFTR conductance was not reversed by returning the temperature to 23°C. The second-site reverent construct, R553M/ ΔF508 CFTR, previously shown to rescue CFTR function in mammalian cells (Teem et al. 1993. Cell. 73:335-346), exhibited a thermal response that was indistinguishable from wild type. Preliminary data suggests that this “thermal instability” that is readily detectable when ΔF508 CFTR is expressed in Xenopus oocytes, reflects an intrinsic structural defect in the channel protein that results in a temperature-sensitive alteration in gating and could potentially trigger the retrieval of surface protein documented in mammalian cells.