title: Neurotensin-induced miR-133α expression regulates neurotensin receptor 1 recycling through its downstream target aftiphilin.
abstract: Neurotensin (NT) triggers signaling in human colonic epithelial cells by activating the G protein-coupled receptor, the neurotensin receptor 1 (NTR1). Activated NTR1 traffics from the plasma membrane to early endosomes, and then recycles. Although sustained NT/NTR1 signaling requires efficient NTR1 recycling, little is known about the regulation of NTR1 recycling. We recently showed that NT/NTR1 signaling increases expression of miR-133α. Herein, we studied the mechanism of NT-regulated miR-133α expression and examined the role of miR-133α in intracellular NTR1 trafficking in human NCM460 colonocytes. We found that NT-induced miR-133α upregulation involves the negative transcription regulator, zinc finger E-box binding homeobox 1. Silencing of miR-133α or overexpression of aftiphilin (AFTPH), a binding target of miR-133α, attenuated NTR1 trafficking to plasma membrane in human colonocytes, without affecting NTR1 internalization. We localized AFTPH to early endosomes and the trans-Golgi network (TGN) in unstimulated human colonic epithelial cells. AFTPH overexpression reduced NTR1 localization in early endosomes and increased expression of proteins related to endosomes and the TGN trafficking pathway. AFTPH overexpression and de-acidification of intracellular vesicles increased NTR1 expression. Our results suggest a novel mechanism of GPCR trafficking in human colonic epithelial cells by which a microRNA, miR-133α regulates NTR1 trafficking through its downstream target AFTPH.
authors.affiliation: Inflammatory Bowel Disease Center, Division of Digestive Diseases, David Geffen School of Medicine, University of California at Los Angeles, California, USA.
authors.forename: Ivy Ka Man
authors.initials: IK
authors.lastname: Law
authors.affiliation: Monash Institute of Pharmaceutical Sciences, ARC Centre of Excellence in Convergent Bio-Nano Science and Technology Parkville, Monash University, Australia.|Department of Anesthesia and Peri-operative Medicine, Monash University, Australia.
authors.forename: Dane
authors.initials: D
authors.lastname: Jensen
authors.affiliation: Monash Institute of Pharmaceutical Sciences, ARC Centre of Excellence in Convergent Bio-Nano Science and Technology Parkville, Monash University, Australia.|Department of Anesthesia and Peri-operative Medicine, Monash University, Australia.|Department of Pharmacology and Therapeutics, University of Melbourne, Australia.
authors.forename: Nigel W
authors.initials: NW
authors.lastname: Bunnett
authors.affiliation: Inflammatory Bowel Disease Center, Division of Digestive Diseases, David Geffen School of Medicine, University of California at Los Angeles, California, USA.
authors.forename: Charalabos
authors.initials: C
authors.lastname: Pothoulakis
mesh_terms: D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D011992:Endosomes / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D006801:Humans; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D009496:Neurotensin / Q000378:metabolism*; D021381:Protein Transport; D018028:Receptors, Neurotensin / Q000235:genetics / Q000378:metabolism*; D021601:trans-Golgi Network / Q000378:metabolism
chemical_list: C538998:AFTPH protein, human; D002352:Carrier Proteins; C548234:MIRN133 microRNA, human; D035683:MicroRNAs; D009419:Nerve Tissue Proteins; D018028:Receptors, Neurotensin; C114036:neurotensin type 1 receptor; D009496:Neurotensin