title: Biphasic Ccl20 regulation by Toll-like receptor 9 through the activation of ERK-AP-1 and non-canonical NF-κB signaling pathways.
abstract: Chemokines play key roles in immune homeostasis and inflammatory response. Considering the role of Ccl20 and Toll-like receptor 9 (TLR9) in gut homeostasis and inflammatory bowel disease (IBD), regulation of Ccl20 by bacterial DNA, the TLR9 ligand, merits in-depth studies.
  
  
  
  We analyzed Ccl20 expression in various epithelial cell (EC) lines by q-PCR and ELISA. In-vivo expression was investigated in isolated murine colonocytes by immunoblotting. Transcriptional regulation of Ccl20 was studied by reporter assays, gene knock-down, electrophoretic mobility shift assay and chromatin immunoprecipitation. Activation of upstream kinases was checked by immunoblotting.
  
  
  
  We showed low levels of Ccl20 expression in mouse colonic ECs, but marked induction by in vivo treatment with bacterial DNA. This corroborated with persistent Ccl20 induction in different EC lines. We found involvement of MAP-kinases during the early hours after stimulation, and a novel AP-1site (-252bp) regulated the expression in colonic ECs. More importantly, mutually exclusive transcriptional regulation by AP-1 (cjun/cfos) and non-canonical NF-κB (RelB/p52) downstream of MEK-ERK and NIK-IKK-α-NF-κB2 (p100) phosphorylation, respectively was responsible for persistent Ccl20 expression in the colonic cells, while canonical NF-κB isoforms played no role.
  
  
  
  Persistent Ccl20 induction by TLR9 in colonic ECs involves early and delayed activation of two independent signaling pathways. This is the first report of non-canonical NF-κB activation and Ccl20 expression in the colonic ECs by TLR9.
  
  
  
  Our study will help to better understand immune regulation by Ccl20 in the intestine and may be exploited for future development of novel therapeutics against IBD.
authors: Division of Clinical Medicine, National Institute of Cholera and Enteric Diseases, Kolkata, India.
authors: Pujarini
authors: P
authors: Dutta
authors: Division of Clinical Medicine, National Institute of Cholera and Enteric Diseases, Kolkata, India.
authors: Atri
authors: A
authors: Ta
authors: Division of Clinical Medicine, National Institute of Cholera and Enteric Diseases, Kolkata, India.
authors: Bhupesh Kumar
authors: BK
authors: Thakur
authors: Division of Clinical Medicine, National Institute of Cholera and Enteric Diseases, Kolkata, India.
authors: Nirmalya
authors: N
authors: Dasgupta
authors: Division of Clinical Medicine, National Institute of Cholera and Enteric Diseases, Kolkata, India. Electronic address: santasabujdas@yahoo.com.
authors: Santasabuj
authors: S
authors: Das
mesh_terms: D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D054418:Chemokine CCL20 / Q000235:genetics* / Q000378:metabolism; D002633:Chemotaxis / Q000187:drug effects; D004269:DNA, Bacterial / Q000494:pharmacology; D003713:Dendritic Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D020895:Enterocytes / Q000187:drug effects / Q000378:metabolism; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007421:Intestine, Small / Q000187:drug effects / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008954:Models, Biological; D053594:Myeloid Differentiation Factor 88 / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D021122:Protein Subunits / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D015398:Signal Transduction; D048029:TNF Receptor-Associated Factor 6 / Q000378:metabolism; D013997:Time Factors; D051217:Toll-Like Receptor 9 / Q000378:metabolism*; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects / Q000235:genetics
chemical_list: C108355:CCL20 protein, human; C493447:CCL20 protein, mouse; D054418:Chemokine CCL20; D004269:DNA, Bacterial; D047908:Intracellular Signaling Peptides and Proteins; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D021122:Protein Subunits; C495371:TLR9 protein, human; D048029:TNF Receptor-Associated Factor 6; C000604754:Tifab protein, human; D051217:Toll-Like Receptor 9; D018808:Transcription Factor AP-1; D048049:Extracellular Signal-Regulated MAP Kinases
keywords: AP-1; Bacterial DNA; Ccl20; IEC; Persistent induction; RelB/P52; TLR9