June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Circadian clock gene Bmal1 is necessary to establish the S-opsin gradient in murine cone photoreceptors
Author Affiliations & Notes
  • Onkar Sawant
    Ophthalmic Research, Cleveland Clinic, Cleveland, Ohio, United States
  • Amanda Horton
    Ophthalmic Research, Cleveland Clinic, Cleveland, Ohio, United States
  • Ivy S Samuels
    Ohio Department of Ophthalmic Research, Louis Stokes Cleveland Veterans Affairs Medical Center, Cleveland, Ohio, United States
  • Sujata Rao
    Ophthalmic Research, Cleveland Clinic, Cleveland, Ohio, United States
  • Footnotes
    Commercial Relationships   Onkar Sawant, None; Amanda Horton, None; Ivy Samuels, None; Sujata Rao, None
  • Footnotes
    Support  This study was supported in part by the NIH-NEI P30 Core Grant (IP30EY025585-01A1) and Unrestricted Grant from The Research to Prevent Blindness, Inc., awarded to the Cole Eye Institute. This research is also funded by R01 EY027077-01 (SR), RPB1503 (SR), E.Matilda Ziegler Foundation - MZF1408 (SR), Knights Templar Eye Foundation award (SR), VA Merit Award i01-BX002754 (IS) and Cleveland Clinic Foundation startup funds (SR).
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 5601. doi:
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    • Get Citation

      Onkar Sawant, Amanda Horton, Ivy S Samuels, Sujata Rao; Circadian clock gene Bmal1 is necessary to establish the S-opsin gradient in murine cone photoreceptors. Invest. Ophthalmol. Vis. Sci. 2017;58(8):5601.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : We recently reported that cyclic light-dark (LD) exposure during early postnatal period is crucial for photoreceptor development and these light dependent developmental processes in the eye could be mediated by thyroid hormones (TH). In the dark reared (DD) animals the retinal circadian clock genes as well as TH components were altered. Based on these observations we hypothesized that the clock genes control TH signaling mediated photoreceptor development.

Methods : To test this hypothesis, photoreceptor specific Crx Cre and Hgrp Cre lines were used to achieve conditional deletion of clock genes Bmal1 and Per2 from developing photoreceptors. ERGs were performed to assay for visual function. Chromatin immunoprecipitation (ChIP) method was used to determine the downstream targets of BMAL1. Immunofluorescence data was confirmed by qRT-PCR and immunoblots.

Results : Deletion of Bmal1 from the cone photoreceptors results in ectopic expression of S-opsin cones in the dorsal retina and a complete loss of the dorsal-ventral S-opsin gradient. Conversely, deletion of Per2, a negative regulator of the circadian clock assembly exhibited the opposite phenotype with an overall decrease in S-opsin expression. Bmal1 mutant animals demonstrated increase in light-adapted b-wave amplitude to 365 nm of light (short wavelength) but decrease in amplitude to 452 nm, 523 nm and 596 nm of lights (medium-long wavelengths). In DD animals we observed a significant decrease in the retinal expression of type II deiodinase (Dio2). Dio2 is required for converting prohormone thyroxin (T4) into biological active triiodothyronine (T3). In the Dio2 null mice we detected ectopic expression of the S-opsin in the dorsal retina which is similar to the Bmal1 mutant phenotype. This suggests an interaction between Bmal1 and Dio2. To validate this interaction we performed ChIP experiments on retinal tissues. Preliminary data indicates that BMAL1 binds to the Dio2 promoter via E-boxes. Accordingly in Bmal1 mutant animals we detect lower levels of Dio2 mRNA and the double heterozygotes for Bmal1 and Dio2 have defects in S-opsin expression while the single heterozygotes are normal.

Conclusions : Based on these findings, we propose a new molecular cascade involving circadian clock gene Bmal1 that establish cone opsin gradient via thyroid activating enzyme Dio2. Our data suggests that Bmal1 is required for cone development and function.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

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