PROJECT ID: PRJNA672101

Data source SRA: PRJNA672101
Description NRL (Neural retina leucine zipper) is a key regulator of the fate determination and gene expression of rod photoreceptors in the retina of many vertebrates. In this study, we observed a unique retinal phenotype in the nrl knockout zebrafish model characterized by reduced rods with shortened outer segments and gradually increased green-cones in adulthood. By tracing and comparing the developmental processes of WT and nrl knockout rods, we found there might be two waves of rod genesis distinguished as nrl-dependent and independent with different starting times. To reveal the underlying cellular and molecular mechanisms, bulk and single-cell RNA-seq were performed. The changes in gene expression and cell proportion of rods and cones agreed well with our histological and immunofluorescence studies. Interestingly, we found that rods exhibited noticeable heterogeneities in the gene expression patterns, and a part of rods in nrl knockout zebrafish misexpressed the green-cone genes, reflecting a hybrid status of rod and green-cone. Furthermore, we identified mafba as a novel regulator of rod genesis, which was responsible for the development of rods in nrl knockout zebrafish. Our study will largely improve the current understanding of the developmental processes and regulatory mechanisms of rods in zebrafish and probably other species, and may facilitate future studies in the fields of retinal development and retinal degeneration.
Key word Neural retina leucine zipper (NRL); rod genesis; retina; double-knockout; green-cone photoreceptors
Publication Liu F, Qin Y, Huang Y, Gao P et al. Rod genesis driven by mafba in an nrl knockout zebrafish model with altered photoreceptor composition and progressive retinal degeneration. PLoS Genet 2022 Mar;18(3):e1009841. PMID: 35245286
Abstract Neural retina leucine zipper (NRL) is an essential gene for the fate determination and differentiation of the precursor cells into rod photoreceptors in mammals. Mutations in NRL are associated with the autosomal recessive enhanced S-cone syndrome and autosomal dominant retinitis pigmentosa. However, the exact role of Nrl in regulating the development and maintenance of photoreceptors in the zebrafish (Danio rerio), a popular animal model used for retinal degeneration and regeneration studies, has not been fully determined. In this study, we generated an nrl knockout zebrafish model via the CRISPR-Cas9 technology and observed a surprising phenotype characterized by a reduced number, but not the total loss, of rods and over-growth of green cones. We discovered two waves of rod genesis, nrl-dependent and -independent at the embryonic and post-embryonic stages, respectively, in zebrafish by monitoring the rod development. Through bulk and single-cell RNA sequencing, we characterized the gene expression profiles of the whole retina and each retinal cell type from the wild type and nrl knockout zebrafish. The over-growth of green cones and mis-expression of green-cone-specific genes in rods in nrl mutants suggested that there are rod/green-cone bipotent precursors, whose fate choice between rod versus green-cone is controlled by nrl. Besides, we identified the mafba gene as a novel regulator of the nrl-independent rod development, based on the cell-type-specific expression patterns and the retinal phenotype of nrl/mafba double-knockout zebrafish. Gene collinearity analysis revealed the evolutionary origin of mafba and suggested that the function of mafba in rod development is specific to modern fishes. Furthermore, the altered photoreceptor composition and abnormal gene expression in nrl mutants caused progressive retinal degeneration and subsequent regeneration. Accordingly, this study revealed a novel function of the mafba gene in rod development and established a working model for the developmental and regulatory mechanisms regarding the rod and green-cone photoreceptors in zebrafish.


Dataset Information

Dataset ID Species Tissue / Organ Experiment type Sample Source dataset ID
1. PRJNA672101 (nrl-KO) Danio rerio retina growth 5mpf, three retinas from nrl-KO zebrafsih SRA: SRR12899703
2. PRJNA672101 (WT) Danio rerio retina baseline 5mpf, three retinas from WT zebrafsih SRA: SRR12899698

Clustering Result

Cluster Cell type Gene id (symbol) Marker class Evidence
1 Bipolar cells ENSDARG00000027236 (rs1a) marker DOI:10.1371/journal.pgen.1009841
2 Bipolar cells ENSDARG00000027236 (rs1a) marker DOI:10.1371/journal.pgen.1009841
2 Bipolar cells ENSDARG00000004358 (gnb3a) marker DOI:10.1371/journal.pgen.1009841
3 Cone ENSDARG00000042529 (gnat2) marker DOI:10.1371/journal.pgen.1009841
3 Cone ENSDARG00000030758 (guca1c) marker DOI:10.1371/journal.pgen.1009841
3 Cone ENSDARG00000070107 (six7) marker DOI:10.1371/journal.pgen.1009841
4 Bipolar cells ENSDARG00000002576 (cabp5a) marker DOI:10.1371/journal.pgen.1009841
5 Rod ENSDARG00000070666 (rhol) marker DOI:10.1371/journal.pgen.1009841
5 Rod ENSDARG00000002193 (rho) marker DOI:10.1371/journal.pgen.1009841
5 Rod ENSDARG00000044199 (gnat1) marker DOI:10.1371/journal.pgen.1009841
5 Rod ENSDARG00000038378 (sagb) marker DOI:10.1371/journal.pgen.1009841
5 Rod ENSDARG00000012125 (cnga1b) marker DOI:10.1371/journal.pgen.1009841
5 Rod ENSDARG00000029898 (cnga1a) marker DOI:10.1371/journal.pgen.1009841
5 Rod ENSDARG00000013393 (guca1b) marker DOI:10.1371/journal.pgen.1009841
5 Rod ENSDARG00000010454 (guca1a) marker DOI:10.1371/journal.pgen.1009841
6 Retinal ganglion cells ENSDARG00000025206 (syt2a) marker DOI:10.1371/journal.pgen.1009841
6 Retinal ganglion cells ENSDARG00000027828 (grin1a) marker DOI:10.1371/journal.pgen.1009841
6 Retinal ganglion cells ENSDARG00000014746 (rbfox1) marker DOI:10.1371/journal.pgen.1009841
10 RPE ENSDARG00000098724 (rgrb) marker DOI:10.1371/journal.pgen.1009841
10 RPE ENSDARG00000008306 (rdh5) marker DOI:10.1371/journal.pgen.1009841
10 RPE ENSDARG00000033760 (pmelb) marker DOI:10.1371/journal.pgen.1009841
10 RPE ENSDARG00000077652 (lratb.2) marker DOI:10.1371/journal.pgen.1009841
11 Horizontal cells ENSDARG00000014018 (lhx1a) marker DOI:10.1371/journal.pgen.1009841
11 Horizontal cells ENSDARG00000020771 (tnr) marker DOI:10.1371/journal.pgen.1009841
12 Microglia ENSDARG00000023188 (lcp1) marker DOI:10.1371/journal.pgen.1009841
12 Microglia ENSDARG00000033735 (ncf1) marker DOI:10.1371/journal.pgen.1009841
13 Muller glia cells ENSDARG00000012671 (inhbaa) marker DOI:10.1371/journal.pgen.1009841
18 Astrocyte ENSDARG00000057598 (s100b) marker DOI:10.1371/journal.pgen.1009841
18 Astrocyte ENSDARG00000040946 (olig2) marker DOI:10.1371/journal.pgen.1009841
18 Astrocyte ENSDARG00000006427 (fabp2) marker DOI:10.1371/journal.pgen.1009841
18 Astrocyte ENSDARG00000042357 (cldnk) marker DOI:10.1371/journal.pgen.1009841

Cluster Cell type Gene id (symbol) Marker class Evidence
1 Bipolar cells ENSDARG00000027236 (rs1a) marker DOI:10.1371/journal.pgen.1009841
1 Bipolar cells ENSDARG00000002576 (cabp5a) marker DOI:10.1371/journal.pgen.1009841
2 Rod ENSDARG00000070666 (rhol) marker DOI:10.1371/journal.pgen.1009841
2 Rod ENSDARG00000002193 (rho) marker DOI:10.1371/journal.pgen.1009841
2 Rod ENSDARG00000044199 (gnat1) marker DOI:10.1371/journal.pgen.1009841
2 Rod ENSDARG00000038378 (sagb) marker DOI:10.1371/journal.pgen.1009841
2 Rod ENSDARG00000012125 (cnga1b) marker DOI:10.1371/journal.pgen.1009841
2 Rod ENSDARG00000029898 (cnga1a) marker DOI:10.1371/journal.pgen.1009841
3 Rod ENSDARG00000044199 (gnat1) marker DOI:10.1371/journal.pgen.1009841
3 Rod ENSDARG00000002193 (rho) marker DOI:10.1371/journal.pgen.1009841
4 Cone ENSDARG00000042529 (gnat2) marker DOI:10.1371/journal.pgen.1009841
4 Cone ENSDARG00000030758 (guca1c) marker DOI:10.1371/journal.pgen.1009841
4 Cone ENSDARG00000070107 (six7) marker DOI:10.1371/journal.pgen.1009841
5 Muller glia cells ENSDARG00000012671 (inhbaa) marker DOI:10.1371/journal.pgen.1009841
6 Retinal ganglion cells ENSDARG00000027828 (grin1a) marker DOI:10.1371/journal.pgen.1009841
7 Horizontal cells ENSDARG00000020771 (tnr) marker DOI:10.1371/journal.pgen.1009841
11 RPE ENSDARG00000098724 (rgrb) marker DOI:10.1371/journal.pgen.1009841
11 RPE ENSDARG00000008306 (rdh5) marker DOI:10.1371/journal.pgen.1009841
11 RPE ENSDARG00000033760 (pmelb) marker DOI:10.1371/journal.pgen.1009841
12 Microglia ENSDARG00000023188 (lcp1) marker DOI:10.1371/journal.pgen.1009841
15 Astrocyte ENSDARG00000042357 (cldnk) marker DOI:10.1371/journal.pgen.1009841
15 Astrocyte ENSDARG00000006427 (fabp2) marker DOI:10.1371/journal.pgen.1009841
15 Astrocyte ENSDARG00000040946 (olig2) marker DOI:10.1371/journal.pgen.1009841