Polarization of specific tropomyosin isoforms in gastrointestinal epithelial cells and their impact on CFTR at the apical surface.
Microfilaments have been reported to be polarized in a number of cell types based both on function and isoform composition. There is evidence that microfilaments are involved in the movement of vesicles and the polarized delivery of proteins to specialized membrane domains. We have investigated the composition of actin microfilaments in gastrointestinal epithelial cells and their role in the delivery of the cystic fibrosis transmembrane conductance regulator (CFTR) into the apical membrane using cultured T84 cells as a model. We identified a specific population of microfilaments containing the tropomyosin (Tm) isoforms Tm5a and/or Tm5b, which are polarized in T84 cell monolayers. Polarization of this microfilament population occurs very rapidly in response to cell-cell and cell-substratum contact and is not inhibited by jasplakinolide, suggesting this involves the movement of intact filaments. Colocalization of Tm5a and/or Tm5b and CFTR was observed in long-term cultures. A reduction in Tm5a and Tm5b expression, induced using antisense oligonucleotides, resulted in an increase in both CFTR surface expression and chloride efflux in response to cAMP stimulation. We conclude that Tm isoforms Tm5a and/or Tm5b mark an apical population of microfilaments that can regulate the insertion and/or retention of CFTR into the plasma membrane.
Dalby-Payne
Jacqueline Rae
JR
Gastroenterology Research, The Children's Hospital at Westmead, Westmead, NSW Z145, Australia.
O'Loughlin
Edward Vincent
EV
Gunning
Peter
P
D008841:Actin Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D000199:Actins / Q000187:drug effects / Q000378:metabolism; D000970:Antineoplastic Agents / Q000494:pharmacology; D002448:Cell Adhesion; D002712:Chlorides / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000378:metabolism*; D047630:Depsipeptides; D004847:Epithelial Cells / Q000378:metabolism*; D041981:Gastrointestinal Tract / Q000378:metabolism*; D006801:Humans; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000494:pharmacology; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport; D014335:Tropomyosin / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D000199:Actins; D000970:Antineoplastic Agents; C505032:CFTR protein, human; D002712:Chlorides; D047630:Depsipeptides; D016376:Oligonucleotides, Antisense; D010456:Peptides, Cyclic; D020033:Protein Isoforms; C498783:TPM3 protein, human; D014335:Tropomyosin; C057531:jasplakinolide; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000242:Cyclic AMP