title: Rescue of defective pancreatic secretion in cystic-fibrosis cells by suppression of a novel isoform of phospholipase C.
abstract: BACKGROUND
  Cystic fibrosis is caused by mutations in the gene encoding an ion-transport protein, the cystic-fibrosis transmembrane conductance regulator (CFTR). Defective secretion of anions is the primary cause of many of the clinical manifestations of cystic fibrosis, including pancreatic insufficiency. We aimed to identify a molecular mechanism from which a new method to circumvent defective pancreatic secretion could be derived.
  
  
  METHODS
  Multiple-human-tissue RT-PCR and semiquantitative RT-PCR analyses were used to examine gene expression. An antisense technique was used in conjunction with radioimmunoassay, Fura-2 spectrofluorometry, immunohistochemistry, and the short-circuit current technique (Ussing chamber) for elucidation of gene function and its application in rescuing defective pancreatic secretion.
  
  
  RESULTS
  We cloned a newly identified gene, NYD-SP27, which has structural similarity to an isoform of phospholipase C. NYD-SP27 was expressed endogenously in human pancreatic-duct cells and upregulated in cystic fibrosis. Suppression of NYD-SP27, by transfection of its antisense into human cystic-fibrosis pancreatic-duct cells, resulted in augmentation of phospholipase-C-coupled calcium-ion release and protein kinase C activity, improvement in the amount of mutated CFTR reaching the plasma membrane, and restoration of cAMP-activated pancreatic anion secretion.
  
  
  CONCLUSIONS
  NYD-SP27 exerts an inhibitory effect on phospholipase-C-coupled processes that depend on calcium ions and protein kinase C, including CFTR trafficking and function. Its upregulation in pancreatic-duct cells may reveal a previously unsuspected defect in cystic fibrosis contributing to pancreatic insufficiency, and thus represents a new target for pharmacological intervention in cystic fibrosis.
authors.affiliation: Epithelial Cell Biology Research Center, Department of Physiology, Faculty of Medicine, Chinese University of Hong Kong, SAR, Shatin, Hong Kong, China.
authors.forename: Hu
authors.initials: H
authors.lastname: Zhu
authors.forename: Jin Xia
authors.initials: JX
authors.lastname: Zhu
authors.forename: Pui Shan
authors.initials: PS
authors.lastname: Lo
authors.forename: Jianmin
authors.initials: J
authors.lastname: Li
authors.forename: Ka Man
authors.initials: KM
authors.lastname: Leung
authors.forename: Dewi Kenneth
authors.initials: DK
authors.lastname: Rowlands
authors.forename: Lai Ling
authors.initials: LL
authors.lastname: Tsang
authors.forename: Mei Kuen
authors.initials: MK
authors.lastname: Yu
authors.forename: Jian Li
authors.initials: JL
authors.lastname: Jiang
authors.forename: Sun Yee
authors.initials: SY
authors.lastname: Lam
authors.forename: Yiu Wa
authors.initials: YW
authors.lastname: Chung
authors.forename: Zuomin
authors.initials: Z
authors.lastname: Zhou
authors.forename: Jiahao
authors.initials: J
authors.lastname: Sha
authors.forename: Hsiao
authors.initials: H
authors.lastname: Chang Chan
mesh_terms: D016375:Antisense Elements (Genetics); D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D003001:Cloning, Molecular; D000242:Cyclic AMP / Q000378:metabolism; D003550:Cystic Fibrosis / Q000235:genetics* / Q000378:metabolism / Q000503:physiopathology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D010188:Exocrine Pancreatic Insufficiency / Q000235:genetics / Q000378:metabolism* / Q000503:physiopathology; D015870:Gene Expression / Q000235:genetics / Q000502:physiology; D006801:Humans; D017136:Ion Transport / Q000235:genetics; D008565:Membrane Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010183:Pancreatic Ducts / Q000166:cytology / Q000502:physiology; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D014162:Transfection / Q000379:methods; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism
chemical_list: D016375:Antisense Elements (Genetics); C505032:CFTR protein, human; D008565:Membrane Proteins; D020033:Protein Isoforms; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000242:Cyclic AMP; D011493:Protein Kinase C; D010738:Type C Phospholipases; D002118:Calcium