S-palmitoylation regulates biogenesis of core glycosylated wild-type and F508del CFTR in a post-ER compartment.
Defects in CFTR (cystic fibrosis transmembrane conductance regulator) maturation are central to the pathogenesis of CF (cystic fibrosis). Palmitoylation serves as a key regulator of maturational processing in other integral membrane proteins, but has not been tested previously for functional effects on CFTR. In the present study, we used metabolic labelling to confirm that wild-type and F508del CFTR are palmitoylated, and show that blocking palmitoylation with the pharmacologic inhibitor 2-BP (2-bromopalmitate) decreases steady-state levels of both wild-type and low temperature-corrected F508del CFTR, disrupts post-ER (endoplasmic reticulum) maturation and reduces ion channel function at the cell surface. PATs (protein acyl transferases) comprise a family of 23 gene products that contain a DHHC motif and mediate palmitoylation. Recombinant expression of specific PATs led to increased levels of CFTR protein and enhanced palmitoylation as judged by Western blot and metabolic labelling. Specifically, we show that DHHC-7 (i) increases steady-state levels of wild-type and F508del CFTR band B, (ii) interacts preferentially with the band B glycoform, and (iii) augments radiolabelling by [3H]palmitic acid. Interestingly, immunofluorescence revealed that DHHC-7 also sequesters the F508del protein to a post-ER (Golgi) compartment. Our findings point to the importance of palmitoylation during wild-type and F508del CFTR trafficking.
Michelle L
ML
McClure
*Gregory Fleming James Cystic Fibrosis Research Center, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
Hui
H
Wen
*Gregory Fleming James Cystic Fibrosis Research Center, University of Alabama at Birmingham, Birmingham, AL 35294, U.S.A.
James
J
Fortenberry
Jeong S
JS
Hong
Eric J
EJ
Sorscher
D000123:Acetyltransferases / Q000145:classification / Q000235:genetics / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000502:physiology*; D005786:Gene Expression Regulation / Q000502:physiology*; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D054878:Lipoylation; D009154:Mutation; D011499:Protein Processing, Post-Translational; D021381:Protein Transport / Q000502:physiology; D011994:Recombinant Proteins
C505032:CFTR protein, human; D047908:Intracellular Signaling Peptides and Proteins; D011994:Recombinant Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000123:Acetyltransferases; C447819:ZDHHC7 protein, human; C062782:protein acyltransferase