PUBLICATION

Eyes shut homolog is important for the maintenance of photoreceptor morphology and visual function in zebrafish

Authors

Messchaert, M., Dona, M., Broekman, S., Peters, T.A., Corral-Serrano, J.C., Slijkerman, R.W.N., van Wijk, E., Collin, R.W.J.

ID

ZDB-PUB-180728-4

Date

2018

Source

PLoS One 13: e0200789 (Journal)

Registered Authors

Collin, Rob, Dona, Margo, Messchaert, Muriel, Peter, Theo, Slijkerman, Ralph, van Wijk, Erwin

Keywords

none

MeSH Terms

RNA/analysis
Genes, Recessive
Vision, Ocular*
Retinitis Pigmentosa/genetics
Rhodopsin/metabolism
Transducin/metabolism
CRISPR-Cas Systems
Genotype
Retina/physiology
Protein Domains
Mutation
DNA Mutational Analysis
Zebrafish
Larva
Retinal Cone Photoreceptor Cells/metabolism
Eye Proteins/genetics*
Eye Proteins/physiology*
Humans
Zebrafish Proteins/genetics*
Zebrafish Proteins/physiology*
Animals
Electroretinography

(all 22)

PubMed

30052645 Full text @ PLoS One

Abstract

Mutations in eyes shut homolog (EYS), a gene predominantly expressed in the photoreceptor cells of the retina, are among the most frequent causes of autosomal recessive (ar) retinitis pigmentosa (RP), a progressive retinal disorder. Due to the absence of EYS in several rodent species and its retina-specific expression, still little is known about the exact function of EYS and the pathogenic mechanism underlying EYS-associated RP. We characterized eys in zebrafish, by RT-PCR analysis on zebrafish eye-derived RNA, which led to the identification of a 8,715 nucleotide coding sequence that is divided over 46 exons. The transcript is predicted to encode a 2,905-aa protein that contains 39 EGF-like domains and five laminin A G-like domains, which overall shows 33% identity with human EYS. To study the function of EYS, we generated a stable eysrmc101/rmc101 mutant zebrafish model using CRISPR/Cas9 technology. The introduced lesion is predicted to result in premature termination of protein synthesis and lead to loss of Eys function. Immunohistochemistry on retinal sections revealed that Eys localizes at the region of the connecting cilium and that both rhodopsin and cone transducin are mislocalized in the absence of Eys. Electroretinogram recordings showed diminished b-wave amplitudes in eysrmc101/rmc101 zebrafish (5 dpf) compared to age- and strain-matched wild-type larvae. In addition, decreased locomotor activity in response to light stimuli was observed in eys mutant larvae. Altogether, our study shows that absence of Eys leads to a disorganized retinal architecture and causes visual dysfunction in zebrafish.

Genes / Markers

Figures

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Expression

Gene	Antibody	Fish	Conditions	Stage	Qualifier	Anatomy	Assay	Figure
eys	Ab1-eys	eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	Not Detected	eye photoreceptor cell	IHC	Fig. 2
eys	Ab1-eys	eys^{rmc101/rmc101} (TL)	standard conditions	Adult	Not Detected	eye photoreceptor cell	IHC	Fig. 2
eys		eys^{rmc101/rmc101} (TL)	standard conditions	Day 5		whole organism	RTPCR	Fig. 1
eys	Ab1-eys	TL	standard conditions	Day 5		eye photoreceptor cell photoreceptor connecting cilium	IHC	Fig. 2
eys	Ab1-eys	TL	standard conditions	Adult		eye photoreceptor cell photoreceptor connecting cilium	IHC	Fig. 2
eys		TL	standard conditions	Day 5		whole organism	RTPCR	Fig. 1
gfap		eys^{rmc101/rmc101} (TL)	standard conditions	Days 45-89		Muller cell	IHC	Fig. S4
gfap		TL	standard conditions	Day 5		Muller cell	IHC	Fig. S4
rho	ab5-rho	eys^{rmc101/rmc101} (TL)	standard conditions	Day 5		eye photoreceptor cell photoreceptor outer segment	IHC	Fig. 2
rho	ab5-rho	eys^{rmc101/rmc101} (TL)	standard conditions	Days 45-89		eye photoreceptor cell photoreceptor outer segment	IHC	Fig. 2

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Phenotype

Fish	Conditions	Stage	Phenotype	Figure
TL	standard conditions	Day 5	Muller cell glial cell activation occurrence, normal	Fig. S4
eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	embryonic retina morphogenesis in camera-type eye decreased process quality, abnormal	Fig. 2
eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	eye photoreceptor cell eys expression absent, abnormal	Fig. 2
eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	eye photoreceptor cell photoreceptor outer segment malformed, abnormal	Fig. 2
eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	optokinetic behavior occurrence, normal	Fig. 3
eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	photoreceptor cell outer segment organization decreased process quality, abnormal	Fig. 2
eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	photoreceptor outer segment layer disorganized, abnormal	Fig. 2
eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	retina detection of light stimulus involved in visual perception decreased process quality, abnormal	Fig. 3
eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	retinal inner nuclear layer thickness, normal	Fig. S3
eys^{rmc101/rmc101} (TL)	standard conditions	Day 5	retinal outer nuclear layer thickness, normal	Fig. S3

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Mutations / Transgenics

Allele	Construct	Type	Affected Genomic Region
rmc101		Small Deletion	eys

Human Disease / Model

Sequence Targeting Reagents

Target	Reagent	Reagent Type
eys	CRISPR4-eys	CRISPR

Fish

Fish
eys^{rmc101/rmc101} (TL)
TL
WT

Antibodies

Name	Type	Antigen Genes	Isotypes	Host Organism
Ab1-cetn	monoclonal		IgG2a	Mouse
Ab1-eys	polyclonal	eys	IgG	Rabbit
Ab2-gnat2	polyclonal		IgG	Rabbit
Ab5-rho	monoclonal	exorh rho		Mouse

Orthology

Engineered Foreign Genes

Mapping

Messchaert et al., 2018 ZDB-PUB-180728-4 PMID:30052645

Eyes shut homolog is important for the maintenance of photoreceptor morphology and visual function in zebrafish

Messchaert et al., 2018

ZDB-PUB-180728-4
PMID:30052645