Section on Molecular Structure and Functional Genomics
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A major emphasis of this group is to move beyond single gene, single technology studies in an attempt to integrate genomics, structure and function and evolutionary perspectives to obtain a wider systems view of the molecular biology of the eye. We have made extensive use of genomics and bioinformatics (largely through NEIBank) to gain new insights into the molecular composition of the eye and to identify sets of specific changes in sequence and expression that distinguish the human (and other primate) eye from those of other mammals, opening the possibility of improving animal models of human eye disease. We are now applying the tools of functional genomics to these discoveries, using techniques including yeast 2-hybrid analysis and knock-out/knock-in mouse models to delineate the roles of key proteins. We also pursue direct biophysical studies of recombinant proteins of interest through collaborations with structural biology labs.NEIBank and Functional Genomics
NEIBank is project for producing, organizing and disseminating genomics resources and expression and sequence data for eye research. The web site is at http://neibank.nei.nih.gov. Collaborators from a wide range of institutions have contributed freshly dissected tissues for cDNA libraries of many eye tissues from many species. These are used for high throughput sequencing. Analyzed data are displayed in the web site, submitted to GenBank and described in peer-reviewed publications. Clones are made available through OpenBiosystems. Eye-related data from other sources are also gathered and displayed through the web site.
Recent additions to NEIBank include:
- Data for several new species of value to the vision research effort, including rhesus monkey. Cow, dog, rabbit, guinea pig, chicken and zebrafish.
- Data for human and mouse lacrimal gland; mouse cornea.
- Data for two human eye-related diseases, keratoconus and pterygium.
- A new browser tool - EyeBrowse. This displays all known expressed sequence tag data for eye tissues aligned against completed genome sequences (currently human, mouse, rat, dog, chicken and zebrafish). It gives a rapid overview of expression patterns for particular genes; shows available clones; identifies variant transcripts; allows display of expression patterns for candidate genes in defined gene linkage loci. http://eyebrowse.cit.nih.gov/
- Human eye disease database. This includes all (as far as possible) genes that have known associations with human eye-related disease. The database also has direct links for each gene to the NCBI SNP database. Disease genes are also displayed on EyeBrowse.
- Library comparison tool: Two libraries (within a species or between species for which homologous gene data available) can be compared side by side with frequency of expression of each gene. This can help identify major differences or similarities among tissues.
- Yeast 2-hybrid libraries for adult mouse lens and retina and for adult human retina and RPE/choroid. These are being used to search for protein interactions involved in age related macular degeneration (AMD) as well as in normal lens and retina function.
- Incorporation of human SAGE and microarray gene expression data for retina and RPE through EyeSAGE, a collaboration with researchers at Duke University.
- cDNA microarrays for eye expressed genes from human and mouse.
The data from NEIBank are a rich source for gene discovery and for insights into species and tissue comparisons. Recent examples of this include the following:
- Discovery of many novel genes, including retbindin, lengsin, opticin/oculoglycan and PDGFD.
- Identification of a novel gene, novel transcripts of known genes and the absence of transcripts for the water channel AQP5 in human keratoconus cornea.
- Identification of novel lacrimal gland genes and a clear illustration of the extensive differences between the human and mouse transcriptional repertoire.
- Analysis of human pterygium shows characteristics of both conjunctiva and corneal epithelium, with makers for the former predominating. Candidate genes for involvement in cell migration have been identified.
- Identification of complete sets of crystallin sequences from several species, with particular emphasis on zebrafish.
- A second
B-crystallin (accompanied by separation of chaperone and crystallin function); gene duplication of AQP0; and a family of five MP19/Lim2 genes in zebrafish.
- Discovery of
Wistow, G. (1995) Molecular Biology and Evolution of Crystallins: Gene Recruitment and Multifunctional Proteins in the Eye Lens. Molecular Biology Intelligence Unit, R.G. Landes Co., Austin TX and Springer, New York.]
Recent observations have also implicated ß - and
We have long standing interests in the genomics, expression, structure and function of crystallins, including collaborative projects for x-ray and NMR structure analyses. Recent work has focused on the analysis of crystallin mutations in cataract and the effects of these mutations as well as differences amongst wild type proteins on stability and solubility. A knock out mouse model is providing new insights into the functional role of
Other proteins with key roles in the lens have been identified through the NEIBank project. Of particular interest are:
- PDGFD (formerly called Iris-expressed growth factor) which appears to have a key role in control of lens cell proliferation and therefore lens growth and is also expressed at high levels in photoreceptors.
- Lengsin is a survivor of an ancient group of class I glutamine synthetases that has lost enzyme activity and has been recruited and re-engineered to a new role in the terminally differentiating fiber cells of the eye lens. Lengsin also shows major modifications during vertebrate evolution that may relate to the different optical requirements of the lens in different species.
Hobby P., Wyatt M.K., Gan W., Bernstein S., Tomarev, S., Slingsby C. and Wistow G. (2000) A Novel Leucine-rich Repeat Protein Expressed in Human Eye: Cloning, Modeling and Chromosomal Localization. Molecular Vision 6, 72-78. PubMed
Wistow, G., Sardarian L., Gan, W. and Wyatt, M.K. (2000) The Human Gene for
Francis, P., Chung, J.-J., Yasui, M., Berry, V., Moore, A., Wyatt, M. K., Wistow, G., Bhattacharya, S. and Agre, P. (2000). Functional impairment of lens aquaporin in two families with dominantly inherited cataracts. Hum Mol Genet 9: 2329-2334 PubMed
Bork, J.M., Peters, L.M., Riazuddin, S., Bernstein, S.L., Ahmed, Z.M., Ness, S.L., Polomeno, R., Ramesh, A., Schloss, M., Srisailpathy, C.R.S., Wayne, S., Bellman, S., Desmukh, D., Ahmed, Z., Khan, S.N., Der Kaloustian, V.M., Li, X.C., Lalwani, A., Riazuddin, S., Bitner-Glindzicz, M., Nance, W.E., Liu, X.-Z., Wistow, G., Smith, R.J.H., Griffith, A.J., Wilcox, E.R., Friedman, T.B., and Morell, R.J. (2001) . Usher syndrome 1D (USH1D) and nonsyndromic recessive deafness DFNB12 are caused by allelic mutations of the novel cadherin-like gene CDH23. Amer. J. Human Genet. 68:26-37. PubMed
Sinha, D., Wyatt, M. K., Sarra, R., Jaworski, C., Slingsby, C., Thaung, C., Pannell, L., Robison, W. G., Favor, J., Lyon, M. and Wistow, G. (2001). A temperature-sensitive mutation of Crygs in the murine Opj cataract. J Biol Chem 276(12): 9308-15. PubMed
Wistow G., (2002) A project for ocular bioinformatics: NEIBank.Mol. Vis. 8:161-163 PubMed
Wistow G, Bernstein SL, Touchman JW, Bouffard G, Wyatt MK, Peterson K, Behal, A., Gao J, Gesuwan P, Smith D. (2002) Grouping and identification of sequence tags (GRIST): Bioinformatics tools for the NEIBank database. Mol Vis. 8:164-170 PubMed
Wistow G, Bernstein SL, Wyatt MK, Behal A, Touchman JW, Bouffard G, Smith D, Peterson K. (2002) NEIBank: Expressed Sequence Tag analysis of adult human lens for the NEIBank project: Over 2000 Unique transcripts, novel genes and splice variants. Mol Vis. 8:171-184 PubMed
Wistow G, Bernstein SL, Ray S, Wyatt MK, Behal A, Touchman JW, Bouffard G, Smith D, Peterson K. (2002) NEIBANK: Expressed sequence tag analysis of adult human iris for the NEIBank Project: Steroid-response factors and similarities with retinal pigment epithelium. Mol Vis. 8:185-195 PubMed
Wistow G, Bernstein SL, Wyatt MK, Ray S, Behal A, Touchman JW, Bouffard G, Smith D, Peterson K. (2002) Expressed sequence tag analysis of human retina for the NEIBank Project: Retbindin, an abundant, novel retinal cDNA and alternative splicing of other retina-preferred gene transcripts. Mol Vis. 8:196-204 PubMed
Wistow G, Bernstein SL, Wyatt MK, Fariss RN, Behal A, Touchman JW, Bouffard G, Smith D, Peterson K. (2002) Expressed sequence tag analysis of human RPE/choroid for the NEIBank Project: Over 6000 non-redundant transcripts, novel genes and splice variants. Mol Vis. 8:205-220 PubMed
Rajini, B., Graham, C., Wistow G. and Sharma Y. (2003). Stability, homodimerization and calcium-binding of a single, variant ß
Tomarev, S. Wistow, G., Raymond, V., Dubois, S., Malyukova, I. (2003). NEIBank: Gene Expression Profile of the Human Trabecular Meshwork. Invest. Ophthalmol. Vis. Sci. 44:2588-2596. PubMed
Bateman, O., Purkiss, A., van Montfort, R., Slingsby, C., Graham, C. and Wistow, G. (2003)The x-ray structure of
Ochrietor,J.D., Moroz, T.P., van Ekeris, L., Clamp, M.F., Jefferson, S.C., deCarvalho, A.C., Fadool, J.M., Wistow, G.J., Muramatsu, T. and Linser, P.J. (2003). Retina expression of 5A11/Basigin-2, a new member of the immunoglobulin superfamily. Invest. Ophthalmol. Vis. Sci. 44:4086-96. PubMed
Wallace, B.A., Wien, F., Miles, A.J., Lees, J.G., Evans, P., Wistow, G.J., and Slingsby, C. (2004). Biomedical applications of synchrotron radiation circular dichroism spectroscopy: Identification of mutant proteins associated with disease and development of a reference database for fold motifs.Faraday Discuss. 126:237 - 243. PubMed
Pompeia C., Hurle, B., Belyantseva, I.A., Noben_Thrauth, K., Beisel, K., Gao, J., Buchoff, P., Wistow, G. and Kachar, B. (2004). Gene Expression Profile of the Mouse Organ of Corti at the Onset of Hearing. Genomics 83:1000-11. PubMed
Ida, H., Boylan, S.A., Weigel, A.L. McBride, Z., Chao, A., Gao, J., Buchoff, P., Wistow, G. and Hjelmeland, L.M. (2004). EST analysis for mouse retina and RPE/choroid cDNA libraries. Mol. Vis. 10:439-444. PubMed
Ahmed, F., Torrado, M., Senatorov, V., Zinovieva, R., Wistow, G. and Tomarev, S. (2004) Gene Expression Profile of the Rat Eye Irido-Corneal Angle: NEIBank Sequence Tag Analysis. Invest. Ophthalmol. Vis. Sci. 45:3081-90 PubMed
Evans, P., Wyatt. M.K., Wistow, G.J., Bateman, O.A., Wallace, B.A. and Slingsby, C. (2004). The P23T cataract mutation causes loss of solubility of folded
Ray, S., Gao, C., Wyatt, M.K., Fariss, R.N., Bundek, A., Zelenka, P. and Wistow, G. (2005). PDGF-D: Tissue-specific expression in the eye and a key role in control of lens epithelial cell proliferation. J Biol Chem. 280(9):8494-502 PubMed
Fan, J., Fariss, R., Purkiss, A., Quinlan, R., Slingsby, C., Wistow, G. and Chepelinsky, A. (2005). Specific Interaction between lens MIP/Aquaporin-0 and two members of the
Rabinowitz, Y., Dong, L. and Wistow, G. (2005). Gene expression profile studies of human Keratoconus cornea for NEIBank: A novel gene and the absence of transcripts for Aquaporin 5. Invest. Ophthalmol. Vis. Sci. 46: 1239-1246. PubMed
Ozyildirim, A.M., Wistow, G. J., Gao, J., Wang, J., Dickinson, D.P., Frierson, H. F. and Laurie, G.W. (2005). The Lacrimal Gland Transcriptome is an Unusually Rich Source of Rare and Poorly Characterized Gene Transcripts. Invest. Ophthalmol. Vis. Sci. 46:1572-80. PubMed
Wistow, G., Wyatt, K., David, L., Gao, C., Bateman, O., Bernstein, S., Tomarev, S., Slingsby, C., Segovia, L and Vihtelic, T. (2005)
Wu, Z., Delaglio, F., Wyatt, K., Wistow, G. and Bax, A. (2005). Solution structure of
Vihtelic, T.S. Fadool, J.M., Gao, J., Thornton, K.A., Hyde, D.R. and Wistow, G. (2005). Expressed sequence tag analysis of zebrafish eye tissues for NEIBank. Mol. Vis. 11:1083-1100. PubMed
Wistow, G. (2006).The NEIBank Project for Ocular Genomics. Data-mining gene expression in human and rodent eye tissues. Prog. Ret. Eye Res. 25(1):43-77. PubMed
Smith, A.A., Wyatt, K., Vacha, J., Vihtelic, T.S., Zigler, J.S., Jr., Wistow, G. and Posner, M. (2006). Gene duplication and separation of functions in
Rickman, C.B., Ebright, J.N., Zavodni, Z.J., Yu, L., Wang, T., Daiger, S.P., Wistow, G., Boon, K. and Hauser, M.A. (2006). Defining the Human Macula Transcriptome and Candidate Retinal Disease Genes using EyeSAGE. Invest. Ophthalmol. Vis. Sci. 47:2305-16. PubMed
Wyatt, K., White, H.E., Wang, L., Bateman, O., Slingsby, C., Orlova, E. and Wistow, G. (2006). Lengsin is a survivor of an ancient family of class I glutamine synthetases re-engineered by evolution for a role in the vertebrate lens Structure. 14:1823-34. PubMed
Nag, N., Ray, S., Wyatt, K., Bogani, D., Favor, J., Lyon, M. and Wistow, G. (2007). The murine No3 cataract is the result of endogenous retroviral insertion in cryge. [Genomics. 2007 Epub ahead of print]. PubMed
|Graeme J. Wistow Ph.D.
|Katherine Peterson Ph.D.||Staff Fellow||Petersonk@nei.nih.gov||301-402-3452|
|Jianguo Fan Ph.D.||Staff Fellowfirstname.lastname@example.org||301-402-4812|
|Nabanita Nag B.Sc.||Technical IRTA Fellowemail@example.com||301-402-5351|
|Sugata Ray Ph.D.||Contractorfirstname.lastname@example.org||301-402-3453|
|M. Keith Wyatt Ph.D.||Contractoremail@example.com||301-402-8649|
|James Gao Ph.D.||Contract Programmerfirstname.lastname@example.org||301-402-6369|
|Patee Buchoff M.Sc.||Contract Programmeremail@example.com||301-402-6365|