June 2021
Volume 62, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2021
Flipping the opsin switch: Cis-regulatory elements required for thyroid hormone-mediated differential expression of the tandemly-replicated long wavelength-sensitive (lws) opsin genes of the zebrafish
Author Affiliations & Notes
  • Deborah L Stenkamp
    Biological Sciences, University of Idaho, Moscow, Idaho, United States
  • Robert Mackin
    Biological Sciences, University of Idaho, Moscow, Idaho, United States
  • Ryuichi Ashino
    Integrated Biosciences, University of Tokyo Graduate School of Frontier Sciences, Kashiwa, Chiba, Japan
  • Shoji Kawamura
    Integrated Biosciences, University of Tokyo Graduate School of Frontier Sciences, Kashiwa, Chiba, Japan
  • Footnotes
    Commercial Relationships   Deborah Stenkamp, None; Robert Mackin, None; Ryuichi Ashino, None; Shoji Kawamura, None
  • Footnotes
    Support  NIH R01 EY012146
Investigative Ophthalmology & Visual Science June 2021, Vol.62, 1678. doi:
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      Deborah L Stenkamp, Robert Mackin, Ryuichi Ashino, Shoji Kawamura; Flipping the opsin switch: Cis-regulatory elements required for thyroid hormone-mediated differential expression of the tandemly-replicated long wavelength-sensitive (lws) opsin genes of the zebrafish. Invest. Ophthalmol. Vis. Sci. 2021;62(8):1678.

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Abstract

Purpose : Human trichromatic color vision requires that the tandemly-replicated LWS/MWS (long- and medium-wavelength sensitive) cone opsin genes are differentially expressed in subsets of cones. We recently reported that the nuclear signaling molecule thyroid hormone (TH) is endogenously involved in differential regulation of the tandemly-replicated lws1/lws2 cone opsin array in zebrafish (Mackin et al., 2019; PNAS). The goal of the current study is to identify cis-regulatory elements residing on the lws locus that are needed for the TH-induced switch from expression of lws2 to lws1.

Methods : We performed stable and transient transgenesis of lws1 and lws2 fluorescent reporter constructs, followed by TH treatment of larvae, confocal imaging of whole eyes, and quantification of cones expressing such reporters (n=9-15 larvae for each condition). Constructs for these assays harbored selected deletions of the lws locus.

Results : Analysis of fluorescent reporter expression within two stable transgenic lines revealed that the same 2.6kb region upstream of lws1 was needed for endogenous patterns of both lws1 and lws2 expression, and for the lws2 to lws1 switch in response to TH. Using transient transgenesis assays, we further dissected this region to identify two, separate ~25bp regions within 0.6kb immediately upstream of lws1. Each of these regions contains a predicted, palindromic TH response element (ppTRE). Deletion of the region containing ppTRE1 resulted in no increase of lws1 in response to TH (# cones expressing lws1 reporter, TH vs. control, p=1.0) and eliminated any expression of lws2. Deletion of the region containing ppTRE2 also resulted in no increase of lws1 in response to TH (p=0.60), and in addition there was no decrease of lws2 in response to TH (p=0.13).

Conclusions : These findings expand our understanding of the mechanisms by which TH differentially regulates the zebrafish lws1/lws2 array. This work contributes to filling in knowledge gaps of visual system development and may also provide insight into how TH regulates other gene arrays including the tandemly replicated LWS/MWS cone opsin array in humans.

This is a 2021 ARVO Annual Meeting abstract.

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