摘要
热休克转录因子4(heat shock transcription factor 4,HSF4)于2002年被首次报道与先天性白内障的发病有关。HSF4有两种剪切形式,HSF4a和HSF4b。在晶状体中只表达HSF4b这种活性形式。
先天性白内障是儿童常见的致盲眼病,晶状体是人体内蛋白质含量最高的组织。蛋白质组学,尤其是差异蛋白质组学在疾病研究方面有着广泛的应用。iTRAQ是一项近年出现的差异蛋白质组学研究技术。在中国家系中首次发现的HSF4突变究竟引起哪些蛋白的表达变化最终导致白内障的发生?本研究拟在人晶状体上皮细胞(human lens cells,HLECs)水平运用iTRAQ技术研究该突变对细胞蛋白表达的影响。
老年性白内障是最为常见的白内障类型,遗传因素被认为和老年性白内障的发生有一定的关系。评价一个基因对白内障发生的影响,应该从一个综合的角度出发来观察,除了与先天性白内障有关,HSF4与老年性白内障的发生是否也存在关系?从这点出发我们收集单纯老年性白内障患者及正常人群血样,进行了相应的研究。
第一部分HSF4b突变对人晶状体细胞株SRA01/04影响的蛋白质组学研究
目的:利用蛋白质组学方法研究HSF4b基因突变(348位核苷酸T突变为C)对人晶状体上皮细胞株SRA01/04的影响。
方法:培养人晶体上皮细胞株SRA01/04,对含人HSF4b基因的质粒进行定点突变,NUucleoBond Xtra Main试剂盒进行质粒抽提,脂质体转染人晶状体上皮细胞(pcDNA3.1-HSF4b,突变的pcDNA3.1-HSF4b,空载体pcDNA3.1),提取四组细胞总蛋白(经过转染的三组细胞和空白细胞),iTRAQ试剂盒标记蛋白,液相色谱串联质谱鉴定方法定量分析并寻找差异蛋白。
结果:共鉴定出104种蛋白,转染突变pcDNA3.1-HSF4b的细胞分别与其他三组细胞相比,共有20种蛋白发生了上调,14种蛋白发生了下调;以转染空载体组为对照,转染突变HSF4b基因组细胞和转染正常HSF4b组细胞分别有12和14种蛋白的表达量发生改变,其中纽带蛋白均有上调,蛋白S100-A13均有下调。
结论:HSF4b突变(348位核苷酸T突变为C)以后,质粒转染人晶状体上皮细胞,能引起部分细胞蛋白表达量的改变,这些蛋白和热休克反应,细胞分化,蛋白质合成,DNA结合等功能有关。
目的:研究HSF4在老年性白内障发生中的作用。
方法:对上海地区150例单纯老年性白内障患者及100例正常人群的HSF4所有外显子进行测序分析。
结果:在正常人群HSF4编码区和外显子及内含子连接区中并没有发现频率大于5%的SNP位点,在白内障患者发现了7个突变的位点,其中5个突变仅在患者中发现(c.1020—25G>A,c.1078A>G,c.1223C>T,c.1286C>T,c.1256+25C>T),另外两个在患者和正常人群中均有发现(c.1019+9C>T,c.1243G>A)。
结论:在老年性白内障患者发现了5个新的HSF4突变位点,提示HSF4突变对老年性白内障的发生有一定的影响。
Heat shock transcription factor 4(HSF4) was reported to be related with congenital cataract for the first time in 2002.HSF4 has two isoforms,HSF4a and HSF4b.In eye lens,HSF4b but not HSF4a is existed.
Congenital cataract is the frequent cause of children blindness.The lens is the highest protein content of any tissue of the body.Proteomics,especially differential proteomics is applied widely in disease studies,iTRAQ is one of the methods appeared recently in differential proteomics.Which proteins are changed by the mutation of HSF4 in a Chinese family which was reported for the first time?We try to find the effect of the mutation on Human lens cells(HLECs) using iTRAQ.
Senile cataract is the most frequent cataract.Genetic factors account partly for senile cataract.It is quite reasonable to evaluate the effect of a gene to cataract on the whole.Besides congenital cataract,HSF4 may play a role in senile cataract.So we collected sporadic senile cataract patients who were free from other diseases and natural population to study.
PartⅠ:The proteome study of HLECs transfected with mutated HSF4b
Purpose To study the proteome changes of HLECs,SRA01/04,which were transfected with mutated HSF4b(T to C transition at nucleotide 348).
Methods Human lens cells(HLECs),SRA01/04,were cultured in vitro. The plasmid,pcDNA3.1-HSF4b was mutated at certain site.Plasmid was amplified on a large scale.HLECs were transfected with plasmids using lipofection(pcDNA3.1-HSF4b,mutant pcDNA3.1-HSF4b,empty plasmid vector pcDNA3.1).The whole cell proteins of four groups(three groups were transfected with different plasmid and one group without transfection) were extracted and were labeled with iTRAQ after digestion.The proteome changes were studied using liquid chromatography and mass spectrometry.
Results One hundred and four proteins were identified on the whole.Comparing with other three groups,the cells transfected with mutant pcDNA3.1-HSF4b had 20 proteins up-regulated and 14 proteins down-regulated.Twelve proteins in the group transfected with mutant pcDNA3.1-HSF4b and 14 proteins in the group transfected with pcDNA3.1-HSF4b were up-or down-regulated compared with transfected with empty vector pcDNA3.1,among which Vinculin was up-regulated and Protein S100-A13 was down-regulated.
Conclusions Some proteins were up-or down-regulated after HLECs were transfected with mutant pcDNA3.1-HSF4b.These changed proteins were related with heat shock response,cell differentiation,protein biosynthesis and DNA binding.
PartⅡ:The study of HSF4 and age related cataract
Purpose To explore the role of HSF4 in the development of senile cataract.
Methods We screened sequence of all exons of the HSF4 gene in 150 senile cataract patients and natural population of 100 from Shanghai.
Results In individuals of natural population,we detected no single-nucleotide polymorphism with frequency higher than 5%in a complete coding region and their exon-intron boundaries.In senile cataract patients,we found 7 sequence variances,among which 5 were present only in 150 senile cataract patients(c.1020-25G＞A,c.1078A＞G,c.1223C＞T,c.1286C＞T,c.1256+25C＞T) and 2 were present both in 150 patients and 100 control subjects(c.1019+9C＞T,c.1243G＞A).
Conclusions We identified 5 new HSF4 gene mutations in 150 senile cataract patients,which indicated that HSF4 mutation account partly for senile cataracts.
引文
[1]. Min JN, Zhang Y, Moskophidis D, Mivechi NF. Unique contribution of heat shock transcription factor 4 in ocular lens development and fiber cell differentiation. [J] Genesis 2004; 40(4): 205-217.
[2]. Graw J. The genetic and molecular basis of congenital eye defects. [J] Nature Reviews Genetics 2003; 4(11): 876-888.
[3]. Bu L, Jin YP, Shi YF, Chu RY, Ban AR, Eiberg H, Andres L, Jiang HS, Zheng GY, Qian MQ, Cui B, Xia Y, Liu J, Hu LD, Zhao GP, Hayden MR, Kong XY. Mutant DNA-binding domain of hsf4 is associated with autosomal dominant lamel lar and marner cataract.[J] Nature Genetics 2002; 31(3): 276-278.
[4]. Tanabe M, Sasai N, Nagata K, Liu XD, Liu PCC, Thiele DJ, Nakai A. The mammalian hsf4 gene generates both an activator and a repressor of heat shock genes by alternative splicing. [J] Journal of Biological Chemistry 1999; 274(39): 27845-27856.
[5]. Nakai A, Tanabe M, Kawazoe Y, Inazawa J, Morimoto RI, Nagata K. Hsf4, a new member of the human heat shock factor family which lacks properties of a transcript ional activator. [J] Molecular and Cellular Biology 1997; 17(1): 469-481.
[6]. Smaoui N, Beltaief O, BenHamed S, M'Rad R, Maazoul F, Ouertani A, Chaabouni H, He jtmancik JF. A homozygous splice mutation in the hsf4 gene is associated with an autosomal recessive congenital cataract. [J] Investigative Ophthalmology & Visual Science 2004; 45(8): 2716-2721.
[7]. Forshew T, Johnson CA, KhaliqS, Pasha S, Willis C, Abbasi R, Tee L, Smith U, Trembath RC, Mehdi SQ, Moore AT, Maher ER. Locus heterogeneity in autosomal recessive congenital cataracts: Linkage to 9q and germline hsf4 mutations. [J] Human Genetics 2005; 117(5): 452-459.
[8]. Ke T, Wang QK, Ji BC, Wang X, Liu P, Zhang XQ, Tang ZH, Ren X, Liu MG. Novel hsf4 mutation causes congenital total white cataract in a chinese family. [J] American Journal of Ophthalmology 2006; 142(2): 298-303.
[9]. Talamas E, Jackson L, Koeberl M, Jackson T, McElwee JL, Hawes NL, Chang B, Jablonski MM, Sidjanin DJ. Early transposable element insertion in intron 9 of the hsf4 gene results in autosomal recessive cataracts in lopll and ldisl mice. [J] Genomics 2006; 88(1): 44-51.
[10]. Mellersh CS, Pettitt L, Forman OP, Vaudin M, Barnett KC. Identification of mutations in hsf4 in dogs of three different breeds with hereditary cataracts. [J] Veterinary Ophthalmology 2006; 9(5): 369-378.
[11]. Fujimoto M, Izu H, Seki K, Fukuda K, Nishida T, Yamada S,Kato K, Yonemura S, Inouye S, Nakai A. Hsf4 is required for normal cell growth and differentiation during mouse lens development. [J] Embo Journal 2004; 23(21): 4297-4306.
[12]. Hammond CJ, Snieder H, Spector TD, Gilbert CE. Genetic and environmental factors in age-related nuclear cataracts in monozygotic and dizygotic twins. [J] New England Journal of Medicine 2000; 342(24): 1786-1790.
[13]. Brookes AJ. The essence of snps. [J] Gene 1999; 234(2): 177-186.
[14]. Iyengar SK, Song DH, Klein BEK, Klein R, Schick JH, Humphrey J, Mi Hard C, Liptak R, Russo K, Jun G, Lee KE, Fijal B,Elston RC. Dissection of genomewide-scan data in extended families reveals a major locus and oligogenic susceptibility for age-related macular degeneration. [J] American Journal of Human Genetics 2004;74(1): 20-39.
[15]. Mori K, Horie-Inoue K, Kohda M, Kawasaki I, Gehlbach PL, Awata T, Yoneya S, Okazaki Y, Inoue S. Association of the htral gene variant with age-related macular degeneration in the japanese population. [J] Journal of Human Genetics 2007; 52(7): 636-641.
[16]. DeAngelis MM, Ji F, Adams S, Morrison MA, Harring AJ, Sweeney MO, Capone A, Miller JW, Dry ja TP, Ott J, Kim IK. Alleles in the htra serine peptidase 1 gene alter the risk of neovascular age-related macular degeneration. [J] Ophthalmology 2008; 115(7): 1209-1215.
[17]. Pirkkala L, Nykanen P, Sistonen L. Roles of the heat shock transcription factors in regulation of the heat shock response and beyond. [J] Faseb Journal 2001; 15(7): 1118-1131.
[18]. Clarke EP, Sanwal BD. Cloning of a human collagen-binding protein, and its homology with rat gp46, chick hsp47 and mouse j6 proteins. [J] Biochimica Et Biophysica Acta 1992; 1129(2): 246-248.
[19]. Ikegawa S, Sudo K, Okui K, Nakamura Y. Isolation, characterization and chromosomal assignment of human colligin-2 gene (cbp2). [J] Cytogenetics and Cell Genetics 1995; 71(2): 182-186.
[20]. Matsuoka S, BallifBA, Smogorzewska A, McDonald ER, Hurov KE, Luo J, Bakalarski CE, Zhao ZM, Solimini N, Lerenthal Y, Shiloh Y, Gygi SP, Elledge SJ. Atm and atr substrate analysis reveals extensive protein networks responsive to DNA damage. [J] Science 2007; 316(5828): 1160-1166.
[21]. RikovaK, Guo A, Zeng Q, Possemato A, Yu J, Haack H, Nardone J, Lee K, Reeves C, Li Y, Hu Y, Tan ZP, Stokes M, Sullivan L, Mitchell J, Wetzel R, MacNeill J, Ren JM, Yuan J, Bakalarski CE, Villen J, Kornhauser JM, Smith B, Li D, Zhou X, Gygi SP, Gu TL, Polakiewicz RD, Rush J, Comb MJ. Global survey of phosphotyrosine signaling identifies oncogenic kinases in lung cancer. [J] Cell 2007; 131(6): 1190-1203.
[22]. Obsilova V, Vecer J, Herman P, Pabianova A, Sulc M, Teisinger J, Boura E, Obsil T. 14-3-3 protein interacts with nuclear localization sequence of forkhead transcription factor foxo4. [J] Biochemistry 2005; 44(34): 11608-11617.
[23]. Davezac N, Tondelier D, Lipecka J, Fanen P, Demaugre F, Debski J, Dadlez M, Schrattenholz A, Cahill MA, Edelman A. Global proteomic approach unmasks involvement of keratins 8 and 18 in the delivery of cystic fibrosis transmembrane conductance regulator (cftr)/delta f508-cftr to the plasma membrane. [J] Proteomics 2004; 4(12): 3833-3844.
[24]. Izawa I, Nishizawa M, Ohtakara K, Ohtsuka K, Inada H, Inagaki M. Identification of mrj, a dnaj/hsp40 family protein, as a keratin 8/18 filament regulatory protein. [J] Journal of Biological Chemistry 2000; 275(44): 34521-34527.
[25]. Inada H, Izawa I, NishizawaM, FujitaE, Kiyono T, Takahashi T, Momoi T, Inagaki M. Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with tradd. [J] Journal of Cell Biology 2001; 155(3): 415-425.
[26]. NishizawaM, Izawa I, Inoko A, Hayashi Y, Nagata K, Yokoyama T, Usukura J, Inagaki M. Identification of trichoplein, a novel keratin filament-binding protein. [J] Journal of Cell Science 2005; 118(5): 1081-1090.
[27]. WadaH, Kito K, Caskey LS, Yeh ETH, Kamitani T. Cleavage of the c-terminus of nedd8 by uch-13. [J] Biochemical and Biophysical Research Communications 1998; 251(3): 688-692.
[28]. .Liu YC, Fallon L, Lashuel HA, Liu ZH, Lansbury PT. The uch-11 gene encodes two opposing enzymatic activities that affect alpha-synuclein degradation and parkinson's disease susceptibility. [J] Cell 2002; 111(2): 209-218.
[29]. Larsen CN, Price JS, Wilkinson KD. Substrate binding and catalysis by ubiquitin c-terminal hydrolases: Identification of two active site residues. [J] Biochemistry 1996; 35(21): 6735-6744.
[30]. Wicki R, Schafer BW, Erne P, Heizmann CW. Characterization of the human and mouse cdnas coding for sl00al3, a new member of the s100 protein family. [J] Biochemical and Biophysical Research Communications 1996; 227(2): 594-599.
[31]. Russo G, Cuccurese M, Monti G, Russo A, Amoresano A, Pucci P, Pietropaolo C. Ribosomal protein 17a binds rna through two distinct rna-binding domains. [J] Biochemical Journal 2005; 385: 289-299.
[32]. Wang H, Kesinger JW, Zhou QC, Wren JD, Martin G, Turner S,Tang YH, Frank MB, Centola M. Identification and characterization of zebrafish ocular formation genes. [J] Genome 2008; 51(3): 222-235.
[33]. Zhang WY, Hawse J, Huang QL, Sheetz N, Miller KM, Horwitz J, Kantorou M. Decreased expression of ribosomal proteins in human age-related cataract. [J] Investigative Ophthalmology & Visual Science 2002; 43(1): 198-204.
[34]. Al Refaii A, Alix JH. Ribosome biogenesis is temperature-dependent and delayed in escherichia coli lacking the chaperones dnak or dnaj.[J] Molecular Microbiology 2009; 71(3): 748-762.
[35]. Tagami H, Ray-Gallet D, Almouzni G, Nakatani Y. Histone h3. 1 and h3. 3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis. [J] Cell 2004; 116(1): 51-61.
[36]. Chow CM, Georgiou A, Szutorisz H, Silva AME, Pombo A, Barahona I, Dargelos E, CanzonettaC, Dillon N. Variant histone h3.3 marks promoters of transcriptionally active genes during mammalian cell division. [J] Embo Reports 2005; 6(4): 354-360.
[37]. Wilmarth PA, Tanner S, Dasari S, Nagalla SR, Riviere MA, Bafna V, Pevzner PA, David LL. Age-related changes in human crystallins determined from comparative analysis of post-translational modifications in young and aged lens: Does deamidation contribute to crystallin insolubility? [J] Journal of Proteome Research 2006; 5(10): 2554-2566.
[38]. Parmley JL, Hurst LD. How do synonymous mutations affect fitness? [J] Bioessays 2007; 29: 515-519.
[1]. Ofarrell PH. High-resolution 2-dimensional electrophoresis of proteins. [J] Journal of Biological Chemistry 1975; 250(10): 4007-4021.
[2]. Miyagi M, Rao KCS. Proteolytic o-18-labeling strategies for quantitative proteoraics. [J] Mass Spectrometry Reviews 2007; 26(1): 121-136.
[3]. Timms JF, Cramer R. Difference gel electrophoresis. [J] Proteomics 2008; 8(23-24): 4886-4897.
[4]. Ong SE, Blagoev B, Kratchmarova I, Kristensen DB, Steen H, Pandey A, Mann M. Stable isotope labeling by amino acids in cell culture, si lac, as a simple and accurate approach to expression proteoraics. [J] Molecular & Cellular Proteomics 2002; 1(5): 376-386.
[5]. Mann M. Functional and quantitative proteomics using si lac. [J] Nature Reviews Molecular Cell Biology 2006; 7(12): 952-958. .
[6]. Gruhler A, Olsen JV, Mohammed S, Mortensen P, Faergeman NJ, Mann M, Jensen ON. Quantitative phosphoproteomics applied to the yeast pheromone signaling pathway. [J] Molecular & Cellular Proteomics 2005; 4(3): 310-327.
[7]. Kratchmarova I, Blagoev B, Haack-Sorensen M, Kassem M, Mann M. Mechanism of divergent growth factor effects in mesenchymal stem cell differentiation. [J] Science 2005; 308(5727): 1472-1477.
[8]. Gygi SP, Rist B, Gerber SA, Turecek F, Gelb MH, Aebersold R. Quantitative analysis of complex protein mixtures using isotope-coded affinity tags. [J] Nature Biotechnology 1999; 17(10):994-999.
[9]. Hunt T, Huang Y, Ross P, Pillai S, Purkayastha S, Pappin D. Protein expression analysis and biomarker identification and quantification using multiplexed isobaric tagging technology - itraq reagents. [J] Molecular & Cellular Proteomics 2004; 3(10):S286-S286.
[10]. Desiderio DM, Kai M. Preparation of stable isotope-incorporated peptide internal standards for field desorption mass-spectrometry quantification of peptides in biologic tissue. [J] Biomedical Mass Spectrometry 1983; 10(8): 471-479.
[11]. Mirgorodskaya 0A, Kozmin YP, Titov MI, Korner R, Sonksen CP, Roepstorff P. Quantitation of peptides and proteins by matrix-assisted laser desorption/ionization mass spectrometry using o-18-labeled internal standards. [J] Rapid Communications in Mass Spectrometry 2000; 14(14): 1226-1232.
[12]. Old WM, Meyer-Arendt K, Aveline-Wolf L, Pierce KG, Mendoza A, Sevinsky JR, Resing KA, Ahn NG. Comparison of label-free methods for quantifying human proteins by shotgun proteomics. [J] Molecular & Cellular Proteomics 2005; 4(10): 1487-1502.
[13]. Bondarenko PV, CheliusD, Shaler TA. Identification and relative quantitation of protein mixtures by enzymatic digestion followed by capillary reversed-phase liquid chromatography-tandem mass spectrometry. [J] Analytical Chemistry 2002; 74(18): 4741-4749.
[14]. Ono M, Shitashige M, Honda K, Isobe T, Kuwabara H, Matsuzuki H, Hirohashi S, Yamada T. Label-free quantitative proteomics using large peptide data sets generated by nanoflow liquid chromatography and mass spectrometry. [J] Molecular & Cellular Proteomics 2006; 5(7): 1338-1347.
[15]. Wiener MC, Sachs JR, Deyanova EG, Yates NA. Differential mass spectrometry: A label-free lc-ms method for finding significant differences in complex peptide and protein mixtures. [J] Analytical Chemistry 2004; 76(20): 6085-6096.
[16]. Koo BS, Lee DY, Ha HS, Kim JC, Kim CW. Comparative analysis of the tear protein expression in blepharitis patients using two-dimensional electrophoresis. [J] Journal of Proteome Research 2005; 4(3): 719-724.
[17]. Grus FH, Augustin AJ. High performance liquid chromatography analysis of tear protein patterns in diabetic and non-diabetic dry-eye patients. [J] European Journal of Ophthalmology 2001; 11(1):19-24.
[18]. Green-Church KB, Nichols KK, Kleinholz NM, Zhang LW, Nichols JJ. Investigation of the human tear film proteome using multiple proteomic approaches. [J] Molecular Vision 2008; 14(55-56):456-470.
[19]. Jonsson M, Markstrom K, Behndig A. Slit-scan tomography evaluation of the anterior chamber and corneal configurations at different ages. [J] Acta Ophthalmologica Scandinavica 2006; 84(1):116-120.
[20]. Tor is CB, Yablonski ME, Wang YL, Camras CB. Aqueous humor dynamics in the aging human eye. [J] American Journal of Ophthalmology 1999; 127(4): 407-412.
[21]. Funding M, Vorum H, Honore B, Nexo E, Ehlers N. Proteomic analysis of aqueous humour from patients with acute corneal rejection. [J] Acta Ophthalmologica Scandinavica 2005; 83(1):31-39.
[22]. Ferreira SM, Lerner SF, Brunzini R, Evelson PA, Llesuy SF. Oxidative stress markers in aqueous humor of glaucoma patients. [J] American Journal of Ophthalmology 2004; 137(1): 62-69.
[23]. Berlau J, Lorenz P, Beck R, Makovitzky J, Schlotzer-Schrehardt U, Thiesen HJ, Guthoff R. Analysis of aqueous humour proteins of eyes with and without pseudoexfoliation syndrome. [J] Graefes Archive for Clinical and Experimental Ophthalmology 2001; 239(10): 743-746.
[24]. Vesaluoma M, Mertaniemi P, Mannonen S, Lehto I, Uusitalo R, Sarna S, Tarkkanen A, Tervo T. Cellular and plasma fibronectin in the aqueous humour of primary open-angle glaucoma, exfoliative glaucoma and cataract patients. [J] Eye 1998; 12: 886-890.
[25]. Schlotzer-Schrehardt U, Lommatzsch J, Kuchle M, Konstas AGP,Naumann GOH. Matrix metalloproteinases and their inhibitors in aqueous humor of patients with pseudoexfoliation syndrome/glaucoma and primary open-angle glaucoma. [J] Investigative Ophthalmology & Visual Science 2003; 44: 1117-1125.
[26]. Hu DN, Ritch R, Liebmann J, Liu YZ, Cheng B, Hu MS. Vascular endothelial growth factor is increased in aqueous humor of glaucomatous eyes. [J] Journal of Glaucoma 2002; 11(5): 406-410.
[27]. Knepper PA, Mayanil CSK, Goossens W, Wertz RD, Holgren C, Ritch R, Allingham RR. Aqueous humor in primary open-angle glaucoma contains an increased level of cd44s. [J] Investigative Ophthalmology & Visual Science 2002; 43(1): 133-139.
[28]. Kim SJ, Kim S, Park J, Lee HK, Park KS, Yu HG, Kim Y. Differential expression of vitreous proteins in proliferative diabetic retinopathy. [J] Current Eye Research 2006; 31(3): 231-240.
[29]. Kim T, Kim SJ, Kim K, Kang UB, Lee C, Park KS, Yu HG, Kim Y. Profiling of vitreous proteomes from proliferative diabetic retinopathy and nondiabetic patients. [J] Proteomics 2007; 7(22): 4203-4215.
[30]. Yu J, Liu F, Cui SJ, Liu Y, Song ZY, Cao H, Chen FE, Wang WJ, Sun T, Wang F. Vitreous proteomic analysis of proliferative vitreoretinopathy. [J] Proteomics 2008; 8(17): 3667-3678.
[31]. Padgaonkar VA, Leverenz VR, Fowler KE, Reddy VN, Giblin FJ. The effects of hyperbaric oxygen on the crystalline of cultured rabbit lenses: A possible catalytic role for copper. [J] Experimental Eye Research 2000; 71(4): 371-383.
[32]. Thompson LJ, Wang F, Proia AD, Peters KG, JarroldB, GreisKD. Proteome analysis of the rat cornea during angiogenesis. [J] Proteomics 2003; 3(11): 2258-2266.
[33]. Srivastava OP, Chandrasekaran D, Pfister RR. Molecular changes in selected epithelial proteins in human keratoconus corneas compared to normal corneas. [J] Molecular Vision 2006; 12(182-86) : 1615-1625.
[34]. Nielsen K, Vorum H, Fagerholm P, Birkenkamp-Demtroder K, Honore B, Ehlers N, Orntoft TF. Proteome profiling of corneal epithelium and identification of marker proteins for keratoconus,a pilot study. [J] Experimental Eye Research 2006; 82(2): 201-209.
[35]. Jurkunas UV, Rawe I, Bitar MS, Zhu C, Harris DL, Colby K, Joyce NC. Decreased expression of peroxiredoxins in fuchs'endothelial dystrophy. [J] Investigative Ophthalmology & Visual Science 2008; 49(7): 2956-2963.
[36]. Jurkunas UV, Bitar MS, Rawe I, Harris DL, Colby K, Joyce NC. Increased clusterin expression in fuchs' endothelial dystrophy. [J] Investigative Ophthalmology & Visual Science 2008; 49(7):2946-2955.
[37]. Jurkunas UV, Bitar M, Rawe I. Colocalization of increased transforming growth factor-beta-induced protein (tgfbip) and clusterin in fuchs endothelial corneal dystrophy. [J] Investigative Ophthalmology & Visual Science 2009; 50(3):1129-1136.
[38]. Colvis CM, Duglas-Tabor Y, Werth KB, Vieira NE, Kowalak JA,Janjani A, Yergey AL, Garland DL. Tracking pathology with proteomics: Identification of in vivo degradation products of alpha b-crystallin. [J] Electrophoresis 2000; 21(11): 2219-2227.
[39]. Weinreb O, van Boekel MAM, Dovrat A, Bloemendal H. Effect of uv-a light on the chaperone-like properties of young and old lens alpha-crystal 1 in. [J] Investigative Ophthalmology & Visual Science 2000; 41(1): 191-198.
[40]. Duncan MK, Xie LK, David LL, Robinson ML, Taube JR, Cui WW, Reneker LW. Ectopic pax6 expression disturbs lens fiber cell differentiation. [J] Investigative Ophthalmology & Visual Science 2004; 45(10): 3589-3598.
[41]. Guest PC, Skynner H, Salim K, Tattersall FD, Knowles MR, Atack JR. Detection of gender differences in rat lens proteins using 2-d-dige. [J] Proteomics 2006; 6(2): 667-676.
[42]. MacCoss MJ, McDonald WH, Saraf A, Sadygov R, Clark JM, Tasto JJ, Gould KL, Wolters D, Washburn M, Weiss A, Clark JI,Yates JR. Shotgun identification of protein modifications from protein complexes and lens tissue. [J] Proceedings of the National Academy of Sciences of the United States of America 2002; 99(12):7900-7905.
[43]. David LL, Wilmarth PA, Rustvold DL, Riviere MA. Global proteomic strategy to quantify oxidized cysteines in human nuclear cataract. [J] Iovs 2005; 46(Suppl. S): 3880.
[44]. Ervin LA, Ball LE, Crouch RK, Schey KL. Phosphorylation and glycosylation of bovine lens mp20. [J] Investigative Ophthalmology & Visual Science 2005; 46(2): 627-635.
[45]. Neal RE, An E, Winn KC, Datiles M, Congdon N, Garland D. Site-specific oxidation of tryptophan and raethionine residues in bbl-crystallin from human lens nucleus: Normal, nuclear cataract,and nuclear/psc cataract. [J] Iovs 2005; 46(Suppl. S): 3889.
[46]. Rousseva L, Paik DC, Dillon J, Ryzhov V, Gail lard ER. Oxindolealanine as a biomarker for cataract development in the human lens. [J] Iovs 2005; 46(Suppl. S): 2897.
[47]. Sergeev YV, Soustov LV, Chelnokov EV, Bityurin NM, Backlund PS, Wingfield PT, Ostrovsky MA, Hejtmancik JF. Increased sensitivity of amino-arm truncated beta a3-crystallin to uv-light-induced photoaggregation. [J] Investigative Ophthalmology & Visual Science 2005; 46(9): 3263-3273.
[48]. Hains PG, Truscott RJW. Post-translational modifications in the nuclear region of young, aged, and cataract human lenses. [J] Journal of Proteome Research 2007; 6: 3935-3943.
[49]. Hains PG, Truscott RJW. Proteome analysis of human foetal, aged and advanced nuclear cataract lenses. [J] Proteomics Clinical Applications 2008; 2(12): 1611-1619.
[50]. Wilmarth PA, Tanner S, Dasari S, Nagalla SR, Riviere MA, Bafna V, Pevzner PA, David LL. Age-related changes in human crystallins determined from comparative analysis of post-translational modifications in young and aged lens: Does deamidation contribute to crystal1 in insolubility? [J] Journal of Proteome Research 2006; 5(10): 2554-2566.
[51]. An E, Lu XN, Flippin J, Devaney JM, Halligan B, Hoffman E, Csaky K, Hathout Y. Secreted proteome profiling in human rpe cell cultures derived from donors with age related macular degeneration and age matched healthy donors. [J] Journal of Proteome Research 2006; 5(10): 2599-2610.
[52]. Shinzato M, Yamashiro Y, Miyara N, Iwamatsu A, Takeuchi K, Umikawa M, Bayarjargal M, Kariya KI, Sawaguchi S. Proteomic analysis of the trabecular meshwork of rats in a steroid-induced ocular hypertension model: Downregulation of type i collagen c-propeptides. [J] Ophthalmic Research 2007; 39: 330-337.
[53]. Tezel G, Yang XJ, Cai J. Proteomic identification of oxidatively modified retinal proteins in a chronic pressure-induced rat model of glaucoma. [J] Investigative Ophthalmology & Visual Science 2005;46(9): 3177-3187.
[54]. Lam TC, Li KK, Lo SCL, Guggenheim JA, To CH. Application of fluorescence difference gel electrophoresis technology in searching for protein biomarkers in chick myopia. [J] Journal of Proteome Research 2007; 6: 4135-4149.
[55]. Bertrand E, Fritsch C, Diether S, Lambrou G, Mueller D, Schaeffel F, Schindler P, Schmid KL, van Oostrum J, Voshol H. Identification of apolipoprotein a-i as a "Stop" Signal for myopia.[J] Molecular & Cellular Proteomics 2006; 5(11): 2158-2166.