Abstract
Purpose: :
Present studies examined the effects of cJun on the cytoplasmic proteins profile in injured corneal epithelium.
Methods: :
cJun was conditionally ablated in corneal epithelium by feeding doxycycline for two weeks to triple transgenic Krt12rtTA/rtTA/tetO-Cre/cJunfloxed/floxed (cJunΔCE/ΔCE) mice. Corneal epithelium debridement (2 mm in diameter) was created in the center of corneas of experimental cJunΔCE/ΔCE (dox-induced) and cJunCEf/CEf (un-induced) triple transgenic mice. Cytoplasmic proteins isolated from injured epithelium healed for 6 and 12 h were subjected to proteomic analysis of multiplex quantitative proteomics method, iTRAQ (isobaric Tagging for Relative and Absolute protein Quantification), coupled with two dimensional nano-liquid chromatography (2D nanoLC) and Q-TOF tandem mass spectrometry (MS/MS), was used to simultaneously determine relative changes in the proteome of corneal epithelium.
Results: :
About 150 proteins were identified by proteomic analysis. Data obtained from un-injured corneas, the ablation of cJun in triple transgenic cJunΔCE/ΔCE mice did not cause any significant changes (greater than 2 folds increase and decrease) in cytoplasmic proteins profile of corneal epithelium as compare to that of triple transgenic cJunCEf/CEf mice. There were little changes of cytoplasmic proteins profile in the injured corneal epithelium healed for 6 h in either cJunΔCE/ΔCE or cJunCEf/CEf mice. After 12 h of debridement, there were significant changes in proteins profile in corneas of both cJunΔCE/ΔCE and cJunCEf/CEf mice. Several proteins exhibited greater than five fold increase in comparison to the uninjured epithelium, serum albumin precursor, major urinary protein 4precursor, lacrimal androgen-binding proteins, prostatic steroid-binding protein, α1-antitrypsin precursor, hemopexin precursor, etc. The patterns of proteins profile changes were independent of the presence of cJun even though in many cases there were lower up-regulation of the proteins in the absence of cJun.
Conclusions: :
cJun is not essential for maintaining the profile of cytoplasmic proteins in uninjured corneal epithelium. Epithelium debridement caused an up-regulation of several cytoplasmic proteins in 12 h. Absence of cJun did not abolish such up-regulation caused by injury. It is possible other members of may compensate the lost function of cJun in corneal epithelium of cJunΔCE/ΔCE mice.
Keywords: wound healing • transgenics/knock-outs • proteomics