bHLH genes and retinal cell fate specification

Abstract

The various cell types in the vertebrate retina arise from a pool of common progenitors. The way that the cell types are specified has been a long-standing issue. Decades of research have yielded a large body of information regarding the involvement of extrinsic factors, and only recently has the function of intrinsic factors begun to emerge. This article reviews recent studies addressing the role of basic helix-loop-helix (bHLH) factors in specifying retinal cell types, with an emphasis on bHLHhierarchies leading to photoreceptor production. Photoreceptor genesis appears to employ two transcriptional pathways: ngn2-->neuroD-->raxL and ath5-->neuroD-->raxL. ngn2 and ath5 function in progenitors, which can potentially develop into different cell types. neuroD represents one of the central steps in photoreceptor specification. Ath5 is also essential for ganglion cell development. It remains to be demonstrated whether a bHLH gene functions as a key player in specifying the other types of retinal cells. Genetic knockout studies have indicated intricate cross-regulation among bHLH genes. Future studies are expected to unveil the mechanism by which bHLH factors network with intrinsic factors and communicate with extrinsic factors to ensure a balanced production of the various types of retinal cells.

Fig. 1

Expression of neuroD in the developing chick retina. (A) Detection of visinin mRNA at E6 with in situ hybridization. (B) Detection of neuroD mRNA at E6 with in situ hybridization. (C,D) Detection of NeuroD protein with antibody at E6 (C) and E9 (D). RPE, retinal pigment epithelium; NE, neuroepithelium; ONL, outer nuclear layer; INL, inner nuclear layer; GCL, ganglion cell layer.

Fig. 2

De novo generation of Visinin⁺ cells from retinal pigment epithelium (RPE) culture under the induction of neuroD. Shown is immunostaining with a monoclonal antibody (7G4). (A) E6 RPE cell culture infected with RCAS–neuroD. (B) The same culture infected with RCAS–GFP.

Fig. 3

Photoreceptor-like morphologies of transdifferentiating cells after reseeding onto poly-ornithine coated cover slips. Shown are Visinin⁺ cells with 3-amino-9-ethylcarbzole (A,B) nitroblue tetrazolium (A,C) or fluorescence (D) as the final substrate of immuno-detection. Arrows: axons. Arrowheads: axonal arboration. Open arrows: inner segment-like structures. Yellow arrow in B points to an oil droplet-like structure in a Hoffman image.

Fig. 4

A schematic diagram of the molecular pathways leading to photoreceptor cells and retinal ganglion cells. The life history of a retinal cell is divided into four developmental phases: proliferation, cessation of mitosis, key step in cell-type specification, and terminal differentiation. ngn2 functions in proliferating progenitor cells, whereas ath5 is involved in postmitotic cells. Both can participate in the genesis of multiple types of cells, including photoreceptor and ganglion cells. ath5 and NSCL1 promote retinal ganglion cell differentiation. The developmental transition from cell proliferation to cell differentiation is a critical window for retinal-cell fate specification, and neuroD may act at this step in specifying the photoreceptor fate. Induction of zebrafish ath5 by shh was proposed by Masai et al. (25) and Stenkamp and Frey (30).