title: miR-106b fine tunes ATG16L1 expression and autophagic activity in intestinal epithelial HCT116 cells.
abstract: BACKGROUND
  The microRNAs (miRNAs) regulate gene expression at the posttranscriptional level. ATG16L1, an essential component for autophagy and a risk gene for Crohn's disease, contains two binding sites in the 3'UTR for miR-17 family, including miRs-20a, -93, -106a, and -106b. The purpose of this study was to assess the effects of these miRNAs on ATG16L1 expression and autophagic activity in HCT116 cells.
  
  
  METHODS
  The functional binding sites in the ATG16L1 3'UTR were evaluated by transfection of pMIR-GLO vectors bearing the wild type or mutant 3'UTR into cells for luciferase reporter assay. The miRNA regulation of ATG16L1 expression was determined by quantitative real-time polymerase chain reaction and Western blot. The miRNA regulation of autophagic activity was evaluated by examining LC3II formation using Western blot and confocal imaging.
  
  
  RESULTS
  Both miR-106a and miR-106b mimics inhibited starvation-induced autophagy. The miR-106b mimic reduced ATG16L1 protein expression. Luciferase reporter assays showed that mutating the binding sequence at the positions 1036 to 1042 abrogated miR-106b regulation of ATG16L1 3'UTR luciferase activity. In addition, miR-106a and miR-106b overexpression inhibited the expression of several other autophagy genes, including ATG12.
  
  
  CONCLUSIONS
  miR-106b targets ATG16L1 and modulates autophagy, partially through the binding site at the 3' end of ATG16L1 3'UTR. miR-106a regulates autophagy, possibly irrelevant to ATG16L1 regulation. Both miR-106a and miR-106b regulate multiple autophagy genes so that they may play an integral role in fine-tuning autophagy.
authors.affiliation: Section of Gastroenterology, Department of Medicine, The University of Chicago, Chicago, Illinois.
authors.forename: Zili
authors.initials: Z
authors.lastname: Zhai
authors.forename: Feng
authors.initials: F
authors.lastname: Wu
authors.forename: Alice Y
authors.initials: AY
authors.lastname: Chuang
authors.forename: John H
authors.initials: JH
authors.lastname: Kwon
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics*; D001343:Autophagy; D000071183:Autophagy-Related Proteins; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
chemical_list: D020413:3' Untranslated Regions; C518959:ATG16L1 protein, human; D000071183:Autophagy-Related Proteins; D002352:Carrier Proteins; C542541:MIRN106 microRNA, human; D035683:MicroRNAs; D012333:RNA, Messenger
references.citation: Nature. 2008 Nov 13;456(7219):264-8
references.pmid: 18849965
references.citation: Mol Cell. 2007 Jul 6;27(1):91-105
references.pmid: 17612493
references.citation: Annu Rev Cell Dev Biol. 2011;27:107-32
references.pmid: 21801009
references.citation: Carcinogenesis. 2012 Nov;33(11):2018-25
references.pmid: 22902544
references.citation: Nat Genet. 2007 Feb;39(2):207-11
references.pmid: 17200669
references.citation: Cell. 2008 Mar 7;132(5):875-86
references.pmid: 18329372
references.citation: Inflamm Bowel Dis. 2013 Mar;19(3):471-80
references.pmid: 23399735
references.citation: PLoS One. 2011 Apr 12;6(4):e18501
references.pmid: 21532750
references.citation: Gastroenterology. 2010 Nov;139(5):1630-41, 1641.e1-2
references.pmid: 20637199
references.citation: Annu Rev Physiol. 2013;75:241-62
references.pmid: 23216414
references.citation: Aging (Albany NY). 2011 Feb;3(2):108-24
references.pmid: 21386132
references.citation: Cell. 2010 Jun 25;141(7):1135-45
references.pmid: 20602997
references.citation: Gut. 2011 Sep;60(9):1229-35
references.pmid: 21406388
references.citation: Cell. 2005 Jan 14;120(1):15-20
references.pmid: 15652477
references.citation: Proc Natl Acad Sci U S A. 2009 Apr 7;106(14):5761-6
references.pmid: 19307576
references.citation: Cell Signal. 2012 Nov;24(11):2179-86
references.pmid: 22781751
references.citation: Methods Mol Biol. 2010;648:193-214
references.pmid: 20700714
references.citation: Blood. 2011 Jul 28;118(4):916-25
references.pmid: 21628417
references.citation: Nucleic Acids Res. 2011 Jan;39(Database issue):D152-7
references.pmid: 21037258
references.citation: Int J Biochem Cell Biol. 2010 Aug;42(8):1348-54
references.pmid: 20227518
references.citation: Autophagy. 2009 Aug;5(6):816-23
references.pmid: 19535919
references.citation: Inflamm Bowel Dis. 2010 Oct;16(10):1729-38
references.pmid: 20848482
references.citation: Biochim Biophys Acta. 2009 Apr;1793(4):664-73
references.pmid: 18706940
references.citation: Inflamm Bowel Dis. 2011 Jan;17(1):241-50
references.pmid: 20812331
references.citation: Gastroenterology. 2008 Nov;135(5):1624-1635.e24
references.pmid: 18835392
references.citation: Nature. 2008 Nov 13;456(7219):259-63
references.pmid: 18849966
references.citation: Gastroenterology. 2009 May;136(5):1689-700
references.pmid: 19422085
references.citation: Arch Biochem Biophys. 2011 Feb 15;506(2):223-35
references.pmid: 21156154
references.citation: J Appl Physiol (1985). 2012 Aug;113(3):459-64
references.pmid: 22700801
references.citation: Nature. 2004 Sep 16;431(7006):350-5
references.pmid: 15372042
references.citation: PLoS One. 2011 Jan 20;6(1):e16221
references.pmid: 21283757
references.citation: Nature. 2011 Jan 20;469(7330):323-35
references.pmid: 21248839
references.citation: Inflamm Bowel Dis. 2007 Aug;13(8):941-6
references.pmid: 17455206
references.citation: Autoimmun Rev. 2012 Mar;11(5):305-14
references.pmid: 20627134