title: Comparative proteomics analysis of degenerative eye lenses of nocturnal rice eel and catfish as compared to diurnal zebrafish.
abstract: OBJECTIVE
  The aim of this study was to determine the lens crystallin diversity of degenerative eyes from the rice eel (Monopterus albus) and walking catfish (Clarias batrachus) as compared to that of zebrafish (Danio rerio) by using comparative proteomics methodologies. We endeavored to investigate the evolution of vertebrate lenses particularly concerning the functional loss of lenses in degenerative eyes of rice eels and catfishes living under an environment of perpetual darkness.
  
  
  METHODS
  Fish lenses were collected and homogenized to extract total soluble proteins. The protein mixtures were separated by one- and two-dimensional gel electrophoresis (1D or 2D gel), plus the newer gel-free shotgun proteomic strategy, followed by in-gel digestion and subjection of the digested protein bands or spots to liquid chromatography coupled with tandem mass spectrometry. The proteomics data were analyzed and compared based on the proteomics databank of zebrafish. The soluble lens protein solutions of three piscine species were also processed by gel-filtration chromatography and 1D sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the comparison and validation of various crystallin families, e.g., α-, β-, and γ-crystallins.
  
  
  RESULTS
  In zebrafish eye lenses, γ-crystallin constituted about 71% and α- and β-crystallins comprised 30% of total lens proteins. In rice eel lenses, very little or almost no α-crystallins were detected and β- and γ-crystallins comprised more than 98% of total lens proteins. In catfish lenses, α- and β-crystallins comprised about 40% and γ-crystallin constitutes 60% of total lens proteins. It was of interest to find that α-crystallin was totally absent in the rice eel in contrast to the presence, albeit with very low amounts, of α-crystallin in similarly nocturnal catfish. The ratio of α-crystallin subunits (αA/αB) was found to be about 20:1 for the catfish lens, in great contrast to the ratio of about 3:1 found for most mammalian lenses. In contrast, β- and γ-crystallins were more abundant in lenses of these three piscine species, similar to mammalian lenses. By proteomics analysis, the most abundant β-crystallins were found to comprise a diverse group of βA1a, βA1-2, βA2a, βA2-2, βA4, βB1, βB2, and βB3 subunit crystallins; the monomeric γ-crystallin class contains γB, γD, γM2, γM3, γM5, γM7, γN-A, γN-B, γS1, and γS2 crystallins.
  
  
  CONCLUSIONS
  In cave or nocturnal animals, the eye is sometimes reduced or eliminated because of adaptation to life in visual darkness. The comparative proteomics analysis of degenerative and normal lenses forms a firm molecular basis to investigate further the evolution of piscine lenses in the future. The total numbers of α-, β-, and γ-crystallins in the three fish species as revealed by the current proteomics methodology clearly indicate the complexity and diversity of crystallin species present in the piscine class of vertebrates. The unexpected finding that α-crystallin is absent in the degenerative eye lenses of rice eel may have some bearing on the chaperone function of α-crystallin in regard to its protective role of preventing protein aggregation in diurnal vertebrate lenses to maintain functional transparency.
authors.affiliation: Institute of Marine Biology, National Sun Yat-Sen University, Kaohsiung, Taiwan.
authors.forename: Yi-Reng
authors.initials: YR
authors.lastname: Lin
authors.forename: Hin-Kiu
authors.initials: HK
authors.lastname: Mok
authors.forename: Yuan-Heng
authors.initials: YH
authors.lastname: Wu
authors.forename: Shih-Shin
authors.initials: SS
authors.lastname: Liang
authors.forename: Chang-Chun
authors.initials: CC
authors.lastname: Hsiao
authors.forename: Chun-Hao
authors.initials: CH
authors.lastname: Huang
authors.forename: Shyh-Horng
authors.initials: SH
authors.lastname: Chiou
mesh_terms: D000818:Animals; D049628:Body Size; D002397:Catfishes / Q000033:anatomy & histology / Q000378:metabolism*; D002457:Cell Extracts; D005591:Chemical Fractionation; D002850:Chromatography, Gel; D002940:Circadian Rhythm; D003459:Crystallins / Q000378:metabolism; D004524:Eels / Q000033:anatomy & histology / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D005136:Eye Proteins / Q000378:metabolism; D029941:Fish Proteins / Q000378:metabolism; D007908:Lens, Crystalline / Q000033:anatomy & histology / Q000378:metabolism* / Q000473:pathology*; D012275:Oryza; D040901:Proteomics / Q000379:methods*; D015027:Zebrafish / Q000033:anatomy & histology / Q000378:metabolism*
chemical_list: D002457:Cell Extracts; D003459:Crystallins; D005136:Eye Proteins; D029941:Fish Proteins
references.citation: Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):5011-5
references.pmid: 8506346
references.citation: Eye (Lond). 1994;8 ( Pt 1):125-9
references.pmid: 8013706
references.citation: J Biol Chem. 1975 May 25;250(10):4007-21
references.pmid: 236308
references.citation: Eur J Biochem. 1994 Oct 1;225(1):1-19
references.pmid: 7925426
references.citation: Monogr Popul Biol. 1971;4:1-219
references.pmid: 5162676
references.citation: J Mol Evol. 1986;24(1-2):121-9
references.pmid: 3104612
references.citation: Proc Natl Acad Sci U S A. 1982 Apr;79(7):2360-4
references.pmid: 6285380
references.citation: Mol Cell Proteomics. 2010 Jul;9(7):1369-82
references.pmid: 20382631
references.citation: Invest Ophthalmol Vis Sci. 2002 Jan;43(1):216-24
references.pmid: 11773034
references.citation: Mol Vis. 2005 Nov 17;11:986-95
references.pmid: 16319818
references.citation: Biochim Biophys Acta. 1986 Jun 23;871(3):324-8
references.pmid: 3707973
references.citation: FEBS Lett. 1985 Feb 11;181(1):1-6
references.pmid: 3972098
references.citation: Dev Genes Evol. 2007 Dec;217(11-12):771-82
references.pmid: 18026750
references.citation: Science. 2000 Jul 28;289(5479):631-3
references.pmid: 10915628
references.citation: Proc Natl Acad Sci U S A. 2006 Feb 21;103(8):2552-7
references.pmid: 16481626
references.citation: Int J Pept Protein Res. 1987 Jul;30(1):108-16
references.pmid: 3499402
references.citation: Amino Acids. 2006 Jun;30(4):369-89
references.pmid: 16583312
references.citation: Clin Chim Acta. 2010 Dec 14;411(23-24):1920-8
references.pmid: 20713032
references.citation: J Pharm Biomed Anal. 2009 Jan 15;49(1):123-8
references.pmid: 19019613
references.citation: Exp Eye Res. 1968 Oct;7(4):603-10
references.pmid: 5716102
references.citation: Mol Vis. 2009 Aug 06;15:1521-9
references.pmid: 19668596
references.citation: J Biochem. 1987 Mar;101(3):751-9
references.pmid: 3110141
references.citation: Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10449-53
references.pmid: 1438232
references.citation: Invest Ophthalmol Vis Sci. 2004 May;45(5):1436-41
references.pmid: 15111599
references.citation: Science. 2001 May 4;292(5518):929-34
references.pmid: 11340206
references.citation: Comp Biochem Physiol Part D Genomics Proteomics. 2008 Jun;3(2):155-71
references.pmid: 20483216
references.citation: FEBS Lett. 1986 May 26;201(1):69-73
references.pmid: 3486783
references.citation: Invest Ophthalmol Vis Sci. 2007 May;48(5):2208-13
references.pmid: 17460281
references.citation: Kaohsiung J Med Sci. 2011 Jan;27(1):1-14
references.pmid: 21329886
references.citation: Ophthalmic Res. 2004 Jan-Feb;36(1):4-24
references.pmid: 15007235
references.citation: Prog Retin Eye Res. 2008 Jan;27(1):89-110
references.pmid: 17962065
references.citation: Proc Natl Acad Sci U S A. 2001 May 22;98(11):6116-20
references.pmid: 11371638
references.citation: Biochim Biophys Acta. 1988 Nov 10;951(1):226-9
references.pmid: 3191133
references.citation: J Exp Biol. 2004 Dec;207(Pt 26):4633-49
references.pmid: 15579559
references.citation: Comp Biochem Physiol C Toxicol Pharmacol. 2004 Jul;138(3):343-50
references.pmid: 15533792
references.citation: Prog Biophys Mol Biol. 1988;51(1):47-76
references.pmid: 3064189
references.citation: Mol Vis. 2008 Apr 25;14:806-14
references.pmid: 18449354
references.citation: Ophthalmic Res. 1986;18(2):90-7
references.pmid: 3737118
references.citation: J Biochem. 1992 Sep;112(3):341-5
references.pmid: 1429521
references.citation: Proteomics. 2002 Jan;2(1):3-10
references.pmid: 11788986
references.citation: Biochemistry. 1991 Jul 9;30(27):6633-6
references.pmid: 2065050
references.citation: Biochim Biophys Acta. 1988 Jun 29;955(1):1-9
references.pmid: 3382668
references.citation: FEBS J. 2006 Feb;273(3):481-90
references.pmid: 16420472
references.citation: Proc Natl Acad Sci U S A. 2002 Jun 11;99(12):7900-5
references.pmid: 12060738
references.citation: FEBS J. 2005 May;272(9):2276-91
references.pmid: 15853812
references.citation: Science. 1969 Sep 5;165(3897):971-81
references.pmid: 17791021
references.citation: Biochem Pharmacol. 2011 Jul 1;82(1):35-42
references.pmid: 21477582
references.citation: Development. 2006 Jul;133(13):2585-93
references.pmid: 16728471
references.citation: Invest Ophthalmol Vis Sci. 1993 Jan;34(1):10-22
references.pmid: 8425816
references.citation: Exp Eye Res. 2004 Aug;79(2):249-61
references.pmid: 15325572
references.citation: Mol Vis. 2005 Sep 20;11:775-91
references.pmid: 16205622
references.citation: Exp Eye Res. 1986 Jul;43(1):77-92
references.pmid: 3732418
references.citation: Comp Biochem Physiol. 1970 May 1;34(1):71-80
references.pmid: 5426561
references.citation: Dev Biol. 1996 Dec 15;180(2):646-63
references.pmid: 8954734
references.citation: Exp Eye Res. 1976 Jan;22(1):1-73
references.pmid: 767125
references.citation: Biochem Mol Biol Int. 1995 Apr;35(4):725-32
references.pmid: 7627123
references.citation: Mol Cell Proteomics. 2008 Oct;7(10):1983-97
references.pmid: 18490355
references.citation: Mol Vis. 2010 Feb 24;16:294-302
references.pmid: 20182557
references.citation: Prog Biophys Mol Biol. 2004 Nov;86(3):407-85
references.pmid: 15302206
references.citation: Invest Ophthalmol Vis Sci. 2002 Jan;43(1):205-15
references.pmid: 11773033
references.citation: Int J Pept Protein Res. 1979 Apr;13(4):409-17
references.pmid: 457334
references.citation: J Hered. 2005 May-Jun;96(3):185-96
references.pmid: 15653557
references.citation: Biochem Biophys Res Commun. 1998 Nov 18;252(2):412-9
references.pmid: 9826544
references.citation: Annu Rev Biochem. 1988;57:479-504
references.pmid: 3052280
references.citation: Science. 1989 Oct 20;246(4928):352-8
references.pmid: 2678474