June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Genetic deletion of M-opsin prevents “M-cone” degeneration in a mouse model of Leber congenital amaurosis
Author Affiliations & Notes
  • HUI XU
    Interdepartmental program in Neuroscience, University of Utah, Salt Lake City, Utah, United States
    Department of Ophthalmology, Baylor College of Medicine, Houston, Texas, United States
  • Nduka Enemchukwu
    Department of Ophthalmology, Baylor College of Medicine, Houston, Texas, United States
  • Yingbin Fu
    Interdepartmental program in Neuroscience, University of Utah, Salt Lake City, Utah, United States
    Department of Ophthalmology, Baylor College of Medicine, Houston, Texas, United States
  • Footnotes
    Commercial Relationships   HUI XU, None; Nduka Enemchukwu, None; Yingbin Fu, None
  • Footnotes
    Support  This work was supported by NIH grants 5R01EY022614, 2P30EY002520, The Sarah Campbell Blaffer Endowment in Ophthalmology, unrestricted grants to the Department of Ophthalmology at Baylor College of Medicine from RPB.
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 4516. doi:
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    • Get Citation

      HUI XU, Nduka Enemchukwu, Yingbin Fu; Genetic deletion of M-opsin prevents “M-cone” degeneration in a mouse model of Leber congenital amaurosis. Invest. Ophthalmol. Vis. Sci. 2017;58(8):4516.

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

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Abstract

Purpose : Mutations in either RPE65 or lecithin-retinol acyltransferase (LRAT) disrupt 11-cis-retinal recycling and cause Leber congenital amaurosis (LCA), the most severe retinal dystrophy in early childhood. In two compatible murine LCA models (Lrat–/– and Rpe65–/–), common pathologic features include: 1) mislocalization of both M- and S-opsins; 2) M-opsin was degraded whereas S-opsin accumulated; 3) S-opsin enriched ventral/central cones degenerate more rapidly than M-opsin enriched dorsal cones. We have shown previously in Lrat–/– model that S-opsin accumulates and aggregates in cones leading to rapid degeneration of ventral/central cones. The objective here is to investigate the mechanism of dorsal cone (“M-cone”) degeneration in a LCA mouse model. Our hypothesis is that M-opsin degradation associated proteasome stress is involved in “M-cone” degeneration.

Methods : We first generated M-opsin (Opn1mw) knockout mice by CRISPR/Cas9. Since cones degenerate rapidly in Lrat–/– mice in the presence of S-opsin (Opn1sw), we bred Opn1mw–/– mice into the Lrat–/–Opn1sw–/– (a LCA model to study “M cones”) background to analyze the preventive effect by M-opsin deletion. The expression and localization of cone opsins (M- and S-opsin) were detected by western blot and immunohistochemistry. Cones were labeled with mouse cone arrestin antibody and counted at 1-, 6-, and 12-month. Proteasome stress was assessed by the accumulation ubiquitinated proteins in cones using an anti-ubiquitin antibody.

Results : M-opsin (Opn1mw) was disrupted by 1-bp insertion at second exon. M-opsin was not detectable in Opn1mw–/– retina by either western blot or Immunohistochemistry. The number of dorsal cones was decreased to 73.87% (N=4, P< 0.01) at 6-month and 54.73% (N=4, P<0.001) at 12-month in Lrat–/–Opn1sw–/– mice compared with Wild-Type(WT), indicating a slow degeneration of dorsal cones. In contrast, the number of dorsal cones of Opn1mw–/–Lrat–/– Opn1sw–/– is similar to that in control Opn1mw–/–Opn1sw–/– and WT mice (i.e. no degeneration). Ubiquitin labeling shows strong ubiquitin signal in dorsal cones of Lrat–/– Opn1sw–/– mice, but it was markedly decreased in dorsal cones of Opn1mw–/–Lrat–/–Opn1sw–/–mice.

Conclusions : Deletion of M-opsin alleviates proteasome stress in Lrat–/–Opn1sw–/– mice, suggesting that M-opsin degradation associated proteasome stress plays a key role in “M-cone” degeneration in Lrat–/–mice.

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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