| Summary: | Cystic fibrosis (CF) is one of the most common autosomal recessive disorders affecting Caucasian populations. Seventy percent of CF alleles produce the protein ΔF508 CFTR, in which a single phenylalanine residue is missing from position 508 of the polypeptide chain. This is a processing mutant: ΔF508 CFTR is functional, but retained in the endoplasmic reticulum (ER) and degraded by the ubiquitin-proteasome pathway. The release of wild-type CFTR from the ER through the secretory pathway requires the formation of a stable form of the protein that is no longer a substrate for ubiquitination ("stable B"), ΔF508 CFTR is unable to reach this stable form and this is the basis for its premature degradation in the ER. The rabbit reticulocyte lysate (RRL) system has been used to measure the ubiquitination of the wild-type and mutant proteins. Following translation at 24<SUP>o</SUP>C, when the incubation temperature is increased to 30<SUP>o</SUP>C, approximately 20% of translated wild-type CFTR reaches a form indistinguishable from stable B (a similar proportion to that observed in the cell), whereas all ΔF508 is ubiquitinated. Interactions with the molecular chaperones hsp90 and hsc70 cease when stable B is reached, indicating they are involved both in the folding, and possibly ubiquitination, of the wild-type protein. ΔF508 CFTR remains associated with hsc70 and hsp90 throughout its lifetime in the RRL. Addition of the hsp90 binding agent geldanamycin to ΔF508 CFTR-expressing RRL post-translationally (but not co-translationally) stabilises approximately 25% of the mutant protein with respect to ubiquitination. Again, the form of the protein produced is indistinguishable from stable B, and stabilisation coincides with its release from the molecular chaperones hsc70 and hsp90. The timing of release from hsc70 indicates a role for this chaperone in particular in the ubiquitination of the protein.
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