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D. E. Borst; Tweety Homolog 3 Expression in the Eye. Invest. Ophthalmol. Vis. Sci. 2008;49(13):780.
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Tweety homolog 3 (Ttyh3) is a member of the Tweety gene family. The NCBI expression profile of ESTs shows that Ttyh3 is expressed in 86% of the tumors in the database including retinoblastoma. Others have shown that Ttyh2 is up-regulated in renal cell carcinoma and colon carcinoma. The genes for both Ttyh2 and Ttyh3 contain 14 exons. Ttyh3 and Ttyh2 are likely to be membrane proteins because they each contain six potential membrane-spanning regions. We have shown that all three tweety homolog genes are expressed in the retina and that Ttyh2 is expressed in the optic nerve. The goal of this study was to determine if Ttyh3 is also expressed in the optic nerve and if it is expressed in the same cell layers of the retina as Ttyh2.
cDNA was synthesized from RNA isolated from mouse optic nerve. Reverse transcriptase PCR (rtPCR) was performed using oligonucleotide primers specific for Ttyh3. The reaction products and known molecular weight markers were separated on polyacrylamide gels. For the in situ hybridization experiments, both sense and antisense digoxigenin-labeled RNA probes that are specific for Ttyh3 and Ttyh2 were synthesized by in vitro transcription. The probe for Ttyh3 is 888 bp long and spans exons 3 to 11. The Ttyh2 probes were used as controls. Probes were hybridized to sections of either rat or mouse retina. After color development with NBT/BCIP, the sections were viewed using a phase contrast microscope.
Ttyh3 was detected in the mouse optic nerve by rtPCR. Both Ttyh3 and Ttyh2 gene transcripts were localized to the ganglion cell layer and the inner nuclear layer of the retina by in situ hybridization. In the optic nerve, Ttyh3 and Ttyh2 are expressed in different cell types. The sense probes for either Ttyh3 or Ttyh2 did not hybridize to the sections, indicating specific detection of these messages by the antisense probes.
rtPCR and in situ hybridization experiments show that Ttyh 3 is expressed in the optic nerve. Others have shown that Ttyh1 is expressed in Muller glia cells. The in situ hybridization experiments described here suggest that these cells may also express Ttyh2 and Ttyh3. Knowledge of Ttyh3's cellular localization will guide us to understanding its role in retinal function in health and disease.
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