Research Summary
The Retina: an Easily Accessible Part of the Central Nervous System

The retina is the most accessible part of the central nervous system for genetic and surgical manipulations. In fact, the retina has been studied as a model system for the brain due to its similar organization, development, and signaling pathways.

Very much involved in retinal diseases is a key component of the rod phototransduction cascade, cGMP-phosphodiesterase (PDE). This enzyme consists of four subunits, two with catalytic activity (PDEalpha and PDEbeta) and two with inhibitory activity (PDEgamma). To examine the function of the inhibitory PDEgamma subunits, Dr. Stephen P. Goff and I removed from the mouse genome the gene that encodes them. This alteration led to an unexpected, dramatic elevation in cGMP and to retinal degeneration resembling human retinitis pigmentosa. A positive participation of PDEgamma in the formation of an active PDE complex is unexpected and unprecedented.

The amount of affected tissue to examine is limited due to the early onset of malformation of photoreceptors in the homozygous mutant. In collaboration with Dr. Victor C. S. Lin (Glaucoma Division), we plan to correct this is to control the expression of this faulty gene by using inducible gene targeting that allows the disruption of a gene active in tissue specific sites at any time in the life of a mouse. In this way the effects of the genetic abnormality can be followed after the photoreceptors have fully developed and are anatomically, physiologically and biochemically normal. An understanding of the early events controlling photoreceptor signaling and degeneration in mice with an induced excision of PDEgamma may help to develop strategies for the prevention or delay of human retinal degenerations. 1 in 3000 individuals is blinded by retinitis pigmentosa, a heterogeneous genetic disorder that featured loss of rod photoreceptor function. One third of us will develop macular degeneration by the 65 years of age.

Gaining temporal and spatial control of gene expression is essential for the elucidation of gene function in the whole organism. The reagents that Dr. Victor C.S. Lin and I plan to develop will also be useful to other laboratories studying the function of specific genes in the retina. An inducible gene targeting system can be used to address several previously unapproachable problems in sensory biology.

Metaphase chromosomes of a genetically manipulated mouse embryonic stem cell line.

Service Activities
Attending Physician, Retina Service, Columbia Medical Center
Diagnosis and management of individuals with retinal vascular diseases such as diabetic retinopathy; retinal degeneration, such as retinitis pigmentosa and macular degeneration; inherited genetic defects affecting the visual system as well as inflammatory eye diseases, such as uveitis; refractive error, such as short-sightedness.

Current Projects

1. Tamoxifen-induced knock-out of a retinal gene
The research project will focus on the generation of tissue- and time- specific modifications in the gene encoding the gamma subunit of cGMP- phosphodiesterase (PDEgamma) by using an inducible knockout system. The entire coding region of the PDEgamma will be flanked by 34-base pair Cre-recombinase recognition sites (loxP) introduced into the gene locus by homologous recombination. After germline transmission, mice Pdeglox1/Pdegtm1 bearing the modified allele will be crossed with mice expressing the Cre-recombinase/steroid hormone receptor fusion protein under the control of the genomic locus of the beta subunit of PDE (Pdeb). The proposed studies will develop tools that will ease the deletion of genes from the genome in specific cells and at specific times. Understanding of the mechanisms controlling photoreceptor signaling and degeneration in Pdeglox1/Pdegtm1 mice may help to develop strategies for the prevention or slowing of human retinal dystrophies.
National Eye Institute
9/2000-8/2008

Selected Publications
1. Farber, D. B., and Tsang, S. H. (2003). Stationary night blindness or progressive retinal degeneration in mice carrying different alleles of PDE gamma, Front Biosci 8, 666-75.

2. Gouras P, Kong J, Tsang SH. Retinal degeneration and RPE transplantation in Rpe65(-/-) mice. Invest Ophthalmol Vis Sci. 2002 Oct;43(10):3307-11.

3. Doi K, Hargitai J, Kong J, Tsang SH, Wheatley M, Chang S, Goff S, Gouras P. Lentiviral transduction of green fluorescent protein in retinal epithelium: evidence of rejection. Vision Res. 2002 Feb;42(4):551-8.

4. Lai CC, Gouras P, Doi K, Tsang SH, Goff SP, Ashton P. Local immunosuppression prolongs survival of RPE xenografts labeled by retroviral gene transfer. Invest Ophthalmol Vis Sci. 2000 Sep;41(10):3134-41.

5. Salchow DJ, Gouras P, Doi K, Goff SP, Schwinger E, Tsang SH. A point mutation (W70A) in the rod PDE-gamma gene desensitizing and delaying murine rod photoreceptors. Invest Ophthalmol Vis Sci. 1999 Dec;40(13):3262-7.

6. Lai CC, Gouras P, Doi K, Lu F, Kjeldbye H, Goff SP, Pawliuk R, Leboulch P, Tsang SH. Tracking RPE transplants labeled by retroviral gene transfer with green fluorescent protein. Invest Ophthalmol Vis Sci. 1999 Aug;40(9):2141-6.