Abstract
Purpose :
A zebrafish emc1-/- line was identified in a mutagenesis screen. Endoplasmic reticulum (ER) membrane protein complex subunit 1 (EMC1) is a protein within a complex responsible for transmembrane protein synthesis and inserting newly folded proteins into the ER membrane. Previous in vitro and in vivo models report Wnt signalling is downstream of Emc1. Here, we sought to investigate cellular and molecular phenotypes in emc1-/- zebrafish.
Methods :
Vision was assessed by optokinetic response (OKR) and visual motor response (VMR). Retinal morphology was assessed by light microscopy. emc1-/- zebrafish bred into Tg(gnat2:eGFP-rab28) transgenic background, were cryosectioned and imaged by confocal microscopy. RNAseq data was analysed using bioinformatic tools and pathway analysis used Panther (www.pantherdb.org). qPCR assessed gene expression. Unpaired t-test and Fisher exact test was used for statistical analysis.
Results :
The OKR to various striped patterns (visual acuity, contrast sensitivity, coloured) is absent in emc1-/- larvae at 131 hours post fertilisation (hpf). Overall VMR locomotor activity is reduced in emc1-/- larvae at 131 hpf (p < 0.0001). emc1-/- have significantly (p<0.04) thinner ganglion cell, inner plexiform and photoreceptor layers. eGFP-rab28 localized almost exclusively to cone outer segments, however, in emc1-/- reduced outer segment labelling was observed. mRNA levels of emc1 were 50% decreased in emc1-/- compared to wildtype (p=0.0056) while emc3 and axin2 were not significantly changed (p=0.1116 and p=0.4481, respectively). The pathway with most significantly decreased genes (log foldchange ≤ -1.5, p<0.05) in emc1-/- was heterotrimeric G-protein signalling pathway-rod outer segment phototransduction (P00028).
Conclusions :
emc1-/- zebrafish larvae have severely impaired vision and display retinal morphology defects. The formation of photoreceptor outer segments is significantly disrupted. Future experiments will assess effects on ocular angiogenesis and the underlying molecular defects.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.