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Robert Mackin, Carmina Gutierrez, Diana Mitchell, Deborah L Stenkamp; The role of thyroid hormone in the differential regulation of tandemly duplicated opsin genes in zebrafish. Invest. Ophthalmol. Vis. Sci. 2016;57(12):576.
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© ARVO (1962-2015); The Authors (2016-present)
Human trichromatic color vision requires a mechanism for differential expression of the tandemly-duplicated LWS/MWS (long- and medium-wavelength sensitive) cone opsin genes, such that they are each expressed in a subset of cones. Our previous published findings have suggested that the trans-acting nuclear signaling molecule retinoic acid (RA) is involved in the differential regulation of tandemly duplicated LWS cone opsins in zebrafish. Here we test the hypotheses that the nuclear signaling molecule thyroid hormone (T3) is also involved in this regulatory activity.
Zebrafish embryos were treated with DMSO (control), 4nM, 20nM, or 100nM T3 from 48 hours post fertilization (hpf) to 96 hpf. Expression of LWS1 and LWS2 was measured by qPCR. To visualize LWS expression, in-situ hybridization using gene specific probes was performed on cryosections. In addition, the transgenic line LWS:PAC(H) was utilized. This line reports LWS1 with GFP and LWS2 with RFP, and these reporters were imaged in whole-mounted embryo eyes with confocal microscopy.
In control embryos, all measures of LWS2 showed widespread and high levels of expression, while all measures of LWS1 showed little or no expression. In embryos treated with 100nM T3, LWS1 expression increased by 10 fold (p=2.84E-07) and was widespread throughout the retina, while LWS2 expression decreased by 5 fold (p=2.16E-07). Embryos treated with 20nM T3 also revealed a significant 9 fold increase in LWS1 (p=2.16E-07) and no significant change in LWS2 (p= 0.98). The effects of 20nM and 100nM T3 were each more robust than the reported effects of 300nM RA. Interestingly, 4nM T3 treatments increased both LWS1 by 7 fold (p=1.12E-06) and LWS2 by 4 fold (p=9.39E-05).
These results significantly extend previous findings that nuclear signaling molecules influence differential regulation of tandemly duplicated cone opsin genes. Our results suggest that the first member of the LWS array (LWS1) responds to exogenous T3 in a dosage-dependent manner. However, LWS2 expression is decreased with 100nM T3 and increased with 4nM T3, suggesting a more complex control mechanism. These findings provide further evidence that the differential regulation of tandemly duplicated cone opsin genes may not rely solely upon stochastic events, but is one in which trans-acting factors can influence differential expression.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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