The role of protein kinase D in neurotensin secretion mediated by protein kinase C-alpha/-delta and Rho/Rho kinase.
Neurotensin (NT) is a gut peptide that plays an important role in gastrointestinal (GI) secretion, motility, and growth as well as the proliferation of NT receptor positive cancers. Secretion of NT is regulated by phorbol ester-sensitive protein kinase C (PKC) isoforms-alpha and -delta and may involve protein kinase D (PKD). The purpose of our present study was: (i) to define the role of PKD in NT release from BON endocrine cells and (ii) to delineate the upstream signaling mechanisms mediating this effect. Here, we demonstrate that small interfering RNA (siRNA) targeted against PKD dramatically inhibited both basal and PMA-stimulated NT secretion; NT release is significantly increased by overexpression of PKD. PKC-alpha and -delta siRNA attenuated PKD activity, whereas overexpression of PKC-alpha and -delta enhanced PKD activity. Rho kinase (ROK) siRNA significantly inhibited NT secretion, whereas overexpression of ROKalpha effectively increased NT release. Rho protein inhibitor C3 dramatically inhibited both NT secretion and PKD activity. In conclusion, our results demonstrate that PKD activation plays a central role in NT peptide secretion; upstream regulators of PKD include PKC-alpha and -delta and Rho/ROK. Importantly, our results identify novel signaling pathways, which culminate in gut peptide release.
Li
Jing
J
Department of Surgery and Sealy Center for Cancer Cell Biology, The University of Texas Medical Branch, 301 University Boulevard, Galveston, TX 77555-0536, USA.
O'Connor
Kathleen L
KL
Hellmich
Mark R
MR
Greeley
George H
GH
Townsend
Courtney M
CM
Evers
B Mark
BM
D015153:Blotting, Western; D002460:Cell Line; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D049452:Green Fluorescent Proteins; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D018613:Microscopy, Confocal; D008954:Models, Biological; D009496:Neurotensin / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D011233:Precipitin Tests; D011493:Protein Kinase C / Q000378:metabolism* / Q000502:physiology*; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000378:metabolism; D011863:Radioimmunoassay; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D020741:rho GTP-Binding Proteins / Q000378:metabolism*; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins; D010455:Peptides; D034741:RNA, Small Interfering; D049452:Green Fluorescent Proteins; D009496:Neurotensin; C089212:protein kinase D; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; C496628:PRKCA protein, human; C497258:PRKCD protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein; D013755:Tetradecanoylphorbol Acetate