Abstract
Purpose :
Melanopsin, the photopigment of pRGCs, modulates retinal function as its genetic deletion leads to disruptions in cone ERG and the baseline and circadian rhythmicity of contrast sensitivity. By analysing retinal gene expression over a circadian time course with RNA-seq, this study aimed to uncover the molecular mechanisms of how melanopsin affects retinal circadian rhythms.
Methods :
Retinas of wild-type (WT) and melanopsin-deficient (Opn4-/-) mice were collected on the first day in constant darkness every 6 hours at circadian times (CT) 4-28 (n=4) and subjected to RNA-seq and qPCR analyses. Circadian rhythmicity and global differences in gene expression were analysed with MetaCycle and DESeq2, respectively. The KEGG database was used to analyse pathway enrichment and the dataset by Siegert et al. (Nat Neurosci 2012) for gene enrichment in retinal cell types.
Results :
WT retinas displayed circadian rhythmicity in 647 genes, and Opn4-/- retinas in 691, while only 178 genes were circadian in both genotypes. Despite these differences, the peak phases of gene expression were remarkably similar between genotypes, with the largest number of genes peaking around CT18. The genes rhythmic in both genotypes showed significant enrichment for dopamine signalling (e.g. Drd1 and Drd4), gap junction and circadian pathways, and cone-associated genes. The genes exclusively circadian in WT retinas were enriched for phototransduction and sphingolipid signalling pathways, and those exclusively rhythmic in Opn4-/- retinas for carbon metabolism. On a global level, 450 genes were differentially expressed between the genotypes and those with higher levels in Opn4-/- were enriched for sphingolipid signalling. The core clock genes Bmal1, Per2 and Cry1 showed circadian rhythmicity in WT, but, interestingly, not in Opn4-/- retinas.
Conclusions :
We demonstrate that retinal circadian genes are associated with dopamine signalling and gap junctions, and are cone-enriched in both WT and Opn4-/- retinas. As demonstrated by Dkhissi-Benyahya et al. (Cell Mol Life Sci 2013), we also find that Opn4-/- retinas lose rhythmicity in core clock gene expression. These findings highlight a critical role for melanopsin in regulating the circadian rhythmicity of the retinal transcriptome and suggest that the circadian disruptions of retinal function in Opn4-/- mice might be a result of disrupted visual transduction and sphingolipid signalling pathways.
This is a 2021 ARVO Annual Meeting abstract.