Abstract
Purpose :
During development, photoreceptor cells (PRCs) are known to arise from columnar neuroepithelium precursor cells and undergo a maturation process to become compartmentalised. Defect in this maturation process can lead to eye diseases. In the past eyes, zebrafish has been shown to be a good model to study eye development and disease. However, a detailed characterisation of PRCs maturation process in this model organism is lacking. In this project, we aimed to establish and characterise in detail the stages of PRC maturation in zebrafish. Next, the role of candidate genes in this PRC maturation process was investigated.
Methods :
To label the plasma membrane of all cells, a zebrafish transgenic line was utilised. Furthermore, a novel zebrafish transgenic line that labels the outer segments of red sensitive PRCs was generated. This transgenic line enabled visualisation and volume quantification of outer segments of red sensitive cones. The use of both transgenic lines in combination with antibody stainings and transmission electron microscopy gave us an insight into the different stages of PRCs maturation. To understand the role of crb2b in the developing retina, a CRISPR mutant was generated and compared to WT.
Results :
From 72 hours post fertilisation (hpf) onwards, subtypes of PRCs in the zebrafish retina exhibit differences in growth rate and morphology of their cell compartments. Additionally, differences in mitochondrial clusters and nuclei positioning were observed during the maturation process. From 72 hpf to 120 hpf, rough endoplasmic reticulum accumulation emerged specifically in rod like PRCs. Changes in chromatin organisation were observed in UV sensitive cones like PRCs from 120 hpf onwards. This showed that a high degree of complexity was observed even within the cone PRC subtypes. Lastly, the role of a candidate gene, crb2b, was examined and the results indicate that loss of Crb2b does not show obvious defects in PRC maturation but leads to degenerative defects in the adult eye.
Conclusions :
This results provided a comprehensive characterisation of six independent PRC maturation stages using the criteria of cell compartmentalisation and growth, organellar distribution and localisation of cell polarity related protein complexes. This defined developmental timeline provides a platform to further study PRC maturation and function.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.