title: Protein expression profiling of lens epithelial cells from Prdx6-depleted mice and their vulnerability to UV radiation exposure.
abstract: Oxidative stress is one of the causative factors in progression and etiology of age-related cataract. Peroxiredoxin 6 (Prdx6), a savior for cells from internal or external environmental stresses, plays a role in cellular signaling by detoxifying reactive oxygen species (ROS) and thereby controlling gene regulation. Using targeted inactivation of the Prdx6 gene, we show that Prdx6-deficient lens epithelial cells (LECs) are more vulnerable to UV-triggered cell death, a major cause of skin disorders including cataractogenesis, and these cells display abnormal protein profiles. PRDX6-depleted LECs showed phenotypic changes and formed lentoid body, a characteristic of terminal cell differentiation and epithelial-mesenchymal transition. Prdx6(-/-) LECs exposed to UV-B showed higher ROS expression and were prone to apoptosis compared with wild-type LECs, underscoring a protective role for Prdx6. Comparative proteomic analysis using fluorescence-based difference gel electrophoresis along with mass spectrometry and database searching revealed a total of 13 proteins that were differentially expressed in Prdx6(-/-) cells. Six proteins were upregulated, whereas expression of seven proteins was decreased compared with Prdx6(+/+) LECs. Among the cytoskeleton-associated proteins that were highly expressed in Prdx6-deficient LECs was tropomyosin (Tm)2beta. Protein blot and real-time PCR validated dramatic increase of Tm2beta and Tm1alpha expression in these cells. Importantly, Prdx6(+/+) LECs showed a similar pattern of Tm2beta protein expression after transforming growth factor (TGF)-beta or H(2)O(2) treatment. An extrinsic supply of PRDX6 could restore Tm2beta expression, demonstrating that PRDX6 may attenuate adverse signaling in cells and thereby maintain cellular homeostasis. Exploring redox-proteomics (Prdx6(-/-)) and characterization and identification of abnormally expressed proteins and their attenuation by PRDX6 delivery should provide a basis for development of novel therapeutic interventions to postpone ROS-mediated abnormal signaling deleterious to cells or tissues.
authors.affiliation: Department of Ophthalmology, Faculty of Medical Science, University of Fukui, 23-3 Shimoaiduki, Matsuoka, Eiheiji, Yoshida-gun, Fukui 910-1193, Japan. kuboe@u-fukui.ac.jp
authors.forename: Eri
authors.initials: E
authors.lastname: Kubo
authors.forename: Nailia
authors.initials: N
authors.lastname: Hasanova
authors.forename: Yukie
authors.initials: Y
authors.lastname: Tanaka
authors.forename: Nigar
authors.initials: N
authors.lastname: Fatma
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Takamura
authors.forename: Dhirendra P
authors.initials: DP
authors.lastname: Singh
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Akagi
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000528:radiation effects; D015153:Blotting, Western; D002478:Cells, Cultured; D019610:Cytoprotection; D004307:Dose-Response Relationship, Radiation; D015180:Electrophoresis, Gel, Two-Dimensional; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology / Q000528:radiation effects*; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000528:radiation effects; D006861:Hydrogen Peroxide / Q000633:toxicity; D007908:Lens, Crystalline / Q000187:drug effects / Q000378:metabolism / Q000473:pathology / Q000528:radiation effects*; D051379:Mice; D018345:Mice, Knockout; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000528:radiation effects; D010449:Peptide Mapping; D054465:Peroxiredoxin VI / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D040901:Proteomics / Q000379:methods*; D012333:RNA, Messenger / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014335:Tropomyosin / Q000378:metabolism; D014466:Ultraviolet Rays
chemical_list: D005136:Eye Proteins; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D011993:Recombinant Fusion Proteins; C490774:Tpm1 protein, mouse; D014335:Tropomyosin; D006861:Hydrogen Peroxide; C515977:PRDX6 protein, human; D054465:Peroxiredoxin VI; C515983:Prdx6 protein, mouse