Purchase this article with an account.
T. Borras, T.V. Morozova, S.L. Heinsohn, R.F. Lyman, T.F. Mackay, R.R. Anholt; Transgenic Flies Overexpressing TIGR/MYOC in the Eye as a New Model for Glaucoma Genetics . Invest. Ophthalmol. Vis. Sci. 2003;44(13):3225.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Purpose: Drosophila melanogaster is a powerful genetic model organism currently utilized as a model for human neurogenerative diseases (1). Our purpose is to characterize transgenic flies with eye specific overexpression of wild-type human TIGR/MYOC, to search for fly eye genes influenced by TIGR/MYOC overexpression, and to identify their human orthologues in the perfused human anterior segment organ culture. Methods: TIGR/MYOC cDNA was cloned into the pUAST vector and the construct introduced into Samarkand w1118 flies. Homozygous transformants were crossed to flies carrying gmr-GAL4 to generate offspring in which GAL4 transactivates eye specific expression of TIGR/MYOC via the UAS promoter in the pUAST vector. RNA extracted from heads of parental and F1 flies was hybridized to Affymetrix GeneChips and data analyzed by one-way-ANOVA. Genes with F1 expression significantly different from the average of the two parental lines (P< 0.01) were retained. Cultured anterior segments from human donors were perfused at 3 µl/min constant flow. At stabilization, eyes were injected through the cornea with recombinant adenoviruses AdhTIG3 (carrying TIGR/MYOC cDNA) (OD) and Ad5CMV.Null (carrying no foreign cDNA) (OS). At 42-72 h, trabecular meshwork (TM) RNA was extracted and analyzed for gene expression by relative quantitative RT-PCR. Results: Overexpression of TIGR/MYOC in the eyes of Drosophila results in distortion of ommatidia accompanied by a periodic fluid discharge from the eye. Microarrays identified altered expression of 50 transcripts, including homologues of aquaporin 4 and cytochrome P-450. Expression of swiss cheese, a neurodegenerative protein, increased 34-fold. Interestingly, its human orthologue, neuropathy target esterase was also up-regulated in the TM of perfused post-mortem human eyes in response to adenovirus mediated TIGR/MYOC overexpression. Conclusions: Proteins affected by TIGR/MYOC expression might have a role in the development of glaucoma. A homologue of a neurodegenerative protein upregulated in transgenic flies is also induced by TIGR/MYOC in the human eye, suggesting it may be associated with regulation of IOP. Our observations establish the Drosophila eye as an advantageous system for the discovery of candidate genes associated with glaucoma. (1) Muqit et al. 2002 Modeling neurodegenerative diseases in Drosophila: a fruitful approach? Nature Rev. Neurosci. 3: 237.
This PDF is available to Subscribers Only