Dataset Information

Cone photoreceptor types in zebrafish are generated by symmetric terminal divisions of dedicated precursors.

ABSTRACT: Proper functioning of sensory systems requires the generation of appropriate numbers and proportions of neuronal subtypes that encode distinct information. Perception of color relies on signals from multiple cone photoreceptor types. In cone-dominated retinas, each cone expresses a single opsin type with peak sensitivity to UV, long (L) (red), medium (M) (green), or short (S) (blue) wavelengths. The modes of cell division generating distinct cone types are unknown. We report here a mechanism whereby zebrafish cone photoreceptors of the same type are produced by symmetric division of dedicated precursors. Transgenic fish in which the thyroid hormone receptor ?2 (tr?2) promoter drives fluorescent protein expression before L-cone precursors themselves are produced permitted tracking of their division in vivo. Every L cone in a local region resulted from the terminal division of an L-cone precursor, suggesting that such divisions contribute significantly to L-cone production. Analysis of the fate of isolated pairs of cones and time-lapse observations suggest that other cone types can also arise by symmetric terminal divisions. Such divisions of dedicated precursors may help to rapidly attain the final numbers and proportions of cone types (L > M, UV > S) in zebrafish larvae. Loss- and gain-of-function experiments show that L-opsin expression requires tr?2 activity before cone differentiation. Ectopic expression of tr?2 after cone differentiation produces cones with mixed opsins. Temporal differences in the onset of tr?2 expression could explain why some species have mixed, and others have pure, cone types.

SUBMITTER: Suzuki SC

PROVIDER: S-EPMC3773785 | biostudies-literature | 2013 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Cone photoreceptor types in zebrafish are generated by symmetric terminal divisions of dedicated precursors.

Suzuki Sachihiro C SC Bleckert Adam A Williams Philip R PR Takechi Masaki M Kawamura Shoji S Wong Rachel O L RO

Proceedings of the National Academy of Sciences of the United States of America 20130826 37

Proper functioning of sensory systems requires the generation of appropriate numbers and proportions of neuronal subtypes that encode distinct information. Perception of color relies on signals from multiple cone photoreceptor types. In cone-dominated retinas, each cone expresses a single opsin type with peak sensitivity to UV, long (L) (red), medium (M) (green), or short (S) (blue) wavelengths. The modes of cell division generating distinct cone types are unknown. We report here a mechanism whe ...[more]

PMID: 23980162

Similar Datasets

Project description:Cone photoreceptors are specialised sensory retinal neurons responsible for photopic vision, colour perception and visual acuity. Retinal degenerative diseases are a heterogeneous group of eye diseases in which the most severe vision loss typically arises from cone photoreceptor dysfunction or degeneration. Establishing a method to purify cone photoreceptors from retinal tissue can accelerate the identification of key molecular determinants that underlie cone photoreceptor development, survival and function. The work herein describes a new method to purify enhanced green fluorescent protein (EGFP)-labelled cone photoreceptors from adult retina of Tg(3.2TαCP:EGFP) zebrafish. Electropherograms confirmed downstream isolation of high-quality RNA with RNA integrity number (RIN) >7.6 and RNA concentration >5.7 ng/µl obtained from both populations. Reverse Transcriptase-PCR (RT-PCR) confirmed that the EGFP-positive cell populations express known genetic markers of cone photoreceptors that were not expressed in the EGFP-negative cell population. This work is an important step towards the identification of cone photoreceptor-enriched genes, protein and signalling networks responsible for their development, survival and function. In addition, this advancement facilitates the identification of novel candidate genes for inherited human blindness. In order to analyse and sort samples by flow cytometry, values for FSC and SSC were displayed in a logarithmic scale, as this is normally the default starting display. This allowed for the identification of different sub-populations of cells present in the retina, which were mixed with unwanted cell debris and cell fragments. Since there were multiple cell populations, different levels of auto-fluorescence were thus successfully detected. It was therefore important to change the strategy and display side scatter and fluorescence characteristics of control and EGFP samples, which ultimately allowed the identification of the extremely well-defined population of EGFP-cone photoreceptors. This improved sorting process minimised RNA degradation. The purified EGFP+ cone photoreceptors represent ~5% of the original dissociated population, which is consistent with humans, wherein the total number of cones (6 million) in the retina is approximately 20 times the one of rods (120 million) (Williamson and Cummins 1983). This work allows high-quality RNA to be obtained from sorted-adult cone photoreceptors. RNA integrity is assessed via 28S and 18S rRNA (Imbeaud et al 2005), and our electropherogram results demonstrate production of high-quality RNA with two clearly visible ribosomal peaks (28S and 18S) from EGFP-sorted cones. In addition, the RNA Integrity Number (RIN), an algorithm for assigning integrity values to RNA based on 28S to 18S rRNA ratios (Sambrook et al 1989; Imbeaud et al 2005; Schroeder et al 2006), had a value of 7.6, higher than the minimum-required 7.0. RNA yields of 5.7 ng/µl were relatively high and sufficient for downstream profiling. RT-PCR confirmed expression of the cone specific gene gnat2, and promoter fragment TαC, but not the retinal pigment epithelium specific gene rpe65 in flow cytometry-sorted GFP-positive photoreceptors (GFP+ cells). rpe65 was neither present in flow cytometry-sorted GFP-negative cones (GFP- cells) as this gene is only expressed in the retinal pigment epithelium (RPE). This study therefore permits the identification of cone photoreceptor-enriched genes, protein and signalling networks responsible for their development, survival and function. In addition, this advancement facilitates the identification of novel candidate genes for inherited human blindness.

Dataset Information

Cone photoreceptor types in zebrafish are generated by symmetric terminal divisions of dedicated precursors.

Publications

Cone photoreceptor types in zebrafish are generated by symmetric terminal divisions of dedicated precursors.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure