dlx and sp6-9 Control Optic Cup Regeneration in a Prototypic Eye

Abstract

Optic cups are a structural feature of diverse eyes, from simple pit eyes to camera eyes of vertebrates and cephalopods. We used the planarian prototypic eye as a model to study the genetic control of optic cup formation and regeneration. We identified two genes encoding transcription factors, sp6-9 and dlx, that were expressed in the eye specifically in the optic cup and not the photoreceptor neurons. RNAi of these genes prevented formation of visible optic cups during regeneration. Planarian regeneration requires an adult proliferative cell population with stem cell-like properties called the neoblasts. We found that optic cup formation occurred only after migration of progressively differentiating progenitor cells from the neoblast population. The eye regeneration defect caused by dlx and sp6-9 RNAi can be explained by a failure to generate these early optic cup progenitors. Dlx and Sp6-9 genes function as a module during the development of diverse animal appendages, including vertebrate and insect limbs. Our work reveals a novel function for this gene pair in the development of a fundamental eye component, and it utilizes these genes to demonstrate a mechanism for total organ regeneration in which extensive cell movement separates new cell specification from organ morphogenesis.

Author Summary

Some invertebrates, such as planarians and Hydra, can regenerate fully after amputations that remove large parts of the body. We investigated how cells in the body of planarians provide new cells for eye regeneration after complete head removal. Planarians possess highly potent regenerative cells (neoblasts) in a compartment inside the worm, and these cells must be present in a body fragment for it to regenerate. We identify a pair of transcription factors, sp6-9 and dlx, that are expressed in the optic cup, and use expression of these genes as markers to demonstrate that lineage restriction of eye cells during regeneration begins within the neoblast compartment. dlx and sp6-9 are essential for formation of optic cup progenitors, and inhibition of these genes with RNA interference results in eyes that lack optic cups after regeneration. During eye development in both flies and vertebrates, progenitors form within a patterned epithelium. Interestingly, planarian eye precursors only aggregate once they have stopped cycling and undergone extensive migration. At this stage they already express markers of the terminally differentiated state. Therefore, we identify a mechanism for eye formation during regeneration and a novel function for a conserved gene pair in eye regeneration.