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Gi Sang Yoon, Nan Gao, Fu-shin X. Yu; Identifying a Novel role of Activating Transcription Factor 3 in TLR5-mediated Protection Against Microbial infection of the Cornea. Invest. Ophthalmol. Vis. Sci. 2012;53(14):3655.
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TLR5 pre-activation in the corneal epithelium by flagellin provides protection against Pseudomonas aeruginosa (PA) keratitis primarily due to reprogramming of gene expression. To understand transcriptional regulation of flagellin-induced reprogramming, we sought to identify transcription factor(s) differentially expressed and its role in regulating flagellin-induced gene expression and the innate immune response in the cornea.
Superarray was used to identify genes differentially expressed in flagellin pretreated human corneal epithelial cells (HCECs) with or without Pseudomonas challenge. The expression of an identified gene, ATF3 in response to flagellin and/or PA in vitro in HCECa and in vivo in B6 mice was assessed by RT-PCR, Western blot and/or immunohistochemistry. The role of ATF3 in flagellin-induced protection was elucidated using ATF3 knockout mice. The genes targeted by ATF3 under different conditions were identified using a cDNA array.
ATF3 was greatly up-regulated in HCECs challenged with PA01 and this up-regulation was further augmented by flagellin pretreatment. A similar pattern of ATF3 expression at the mRNA and protein levels was observed in the corneal epithelia of B6 mice. Strikingly, ATF3-/- mice were more susceptible to Pseudomonas infection and voided flagellin-induced protection. While only two genes were detected with altered expression in untreated corneal epithelia of ATF3-/- mice, a large number of genes were identified as ATF3 target genes in infected corneas with or without flagellin pretreatment.
ATF3 is an important effector in mediating the expression of innate defense and cytoprotective genes in corneal epithelial cells. Further analysis of gene expression in flagellin pretreated and/or PA01 infected corneas will reveal ATF3 target genes which are required for corneal innate defense against microbial infection.
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