With a long-term goal to better understand the molecular mechanisms of epithelial innate immune responses to pathogens, we will investigate innate immunity in cholangiocytes (epithelial cells lining the biliary tract) in response to Cryptosporidium parvum, an NIAID Category B Priority Pathogen that causes both intestinal and biliary disease in humans. Using an in vitro model of human biliary cryptosporidiosis established by us, we have demonstrated that: (i) two Toll-like receptors (TLRs), TLR2 and TLR4, and an associated intracellular signaling pathway (NF-kappaB activation) play a central role in C. parvum recognition and induction of innate immune responses (e.g., release of cytokines/ chemokines) in cholangiocytes; (ii) C. parvum infection alters ? cholangiocyte expression of specific endogenous microRNAs (miRNAs), a newly identified class of small regulatory RNAs important in post-transcriptional gene regulation; and (iii) TLR/NF-kappaB signals are involved in C. parvum-induced cholangiocyte miRNA expression. Thus, we will test the CENTRAL HYPOTHESIS that epithelial innate immunity in response to C. parvum infection involves TLR-mediated pathogen recognition and subsequent alteration of miRNA-mediated post-transcriptional gene regulation. In our three integrated SPECIFIC AIMS, we will test the hypotheses that: (i) C. parvum activation of host-cell TLRs alters cholangiocyte expression of endogenous miRNAs; (ii) C. parvum-induced miRNA expression in cholangiocytes is mediated by TLR activation of the nuclear transcription factor, NF-kappaB; and (iii) C. parvum-induced alterations in miRNA expression influence associated post-transcriptional gene regulation and contribute to cholangiocyte innate immune responses. Innovative aspects of the application include novel methodologies for miRNA analysis, an in vitro model of human biliary cryptosporidiosis, and new concepts (regulation of miRNA expression by transcription factors and miRNA-mediated post-transcriptional gene regulation in epithelial innate immunity). These studies will address a fundamental question related to epithelial innate immunity to a Category B Pathogen; specifically, what is the role of miRNAs in TLRmediated cholangiocyte innate immune responses to C. parvum? This NEW CONCEPT is likely relevant to innate and adaptive immunity in general and our results could also provide a rational basis for the design and implementation of new therapeutic strategies for microbial infection. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
7R01AI071321-02
Application #
7414665
Study Section
Special Emphasis Panel (ZAI1-AR-I (M2))
Program Officer
Wali, Tonu M
Project Start
2006-07-01
Project End
2010-06-30
Budget Start
2007-04-07
Budget End
2007-06-30
Support Year
2
Fiscal Year
2006
Total Cost
$155,014
Indirect Cost
Name
Creighton University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
053309332
City
Omaha
State
NE
Country
United States
Zip Code
68178
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Zhou, Rui; Gong, Ai-Yu; Chen, Dongqing et al. (2013) Histone deacetylases and NF-kB signaling coordinate expression of CX3CL1 in epithelial cells in response to microbial challenge by suppressing miR-424 and miR-503. PLoS One 8:e65153
Zhao, Jian; Gong, Ai-Yu; Zhou, Rui et al. (2012) Downregulation of PCAF by miR-181a/b provides feedback regulation to TNF-ýý-induced transcription of proinflammatory genes in liver epithelial cells. J Immunol 188:1266-74
Zhou, Rui; Gong, Ai-Yu; Eischeid, Alex N et al. (2012) miR-27b targets KSRP to coordinate TLR4-mediated epithelial defense against Cryptosporidium parvum infection. PLoS Pathog 8:e1002702
Zhou, Rui; Li, Xiaoqing; Hu, Guoku et al. (2012) miR-16 targets transcriptional corepressor SMRT and modulates NF-kappaB-regulated transactivation of interleukin-8 gene. PLoS One 7:e30772
Gong, Ai-Yu; Hu, Guoku; Zhou, Rui et al. (2011) MicroRNA-221 controls expression of intercellular adhesion molecule-1 in epithelial cells in response to Cryptosporidium parvum infection. Int J Parasitol 41:397-403
Zhou, Rui; O'Hara, Steven P; Chen, Xian-Ming (2011) MicroRNA regulation of innate immune responses in epithelial cells. Cell Mol Immunol 8:371-9
Gong, Ai-Yu; Zhou, Rui; Hu, Guoku et al. (2010) Cryptosporidium parvum induces B7-H1 expression in cholangiocytes by down-regulating microRNA-513. J Infect Dis 201:160-9
O'Hara, Steven P; Splinter, Patrick L; Gajdos, Gabriella B et al. (2010) NFkappaB p50-CCAAT/enhancer-binding protein beta (C/EBPbeta)-mediated transcriptional repression of microRNA let-7i following microbial infection. J Biol Chem 285:216-25
Hu, Guoku; Zhou, Rui; Liu, Jun et al. (2010) MicroRNA-98 and let-7 regulate expression of suppressor of cytokine signaling 4 in biliary epithelial cells in response to Cryptosporidium parvum infection. J Infect Dis 202:125-35

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