Analysis Of Hes1 Function In Hedgehog Signaling During Mouse Retinal Development

Abstract

Purpose: : The mature vertebrate neural retina consists of six major types of neuronal and glial cells and is an excellent system for studying neuronal differentiation. Accumulating evidence indicates that proliferation and cell fate specification of neural progenitor cells are regulated by both cell-intrinsic factors and cell-extrinsic cues. Among the known growth factors, the Hedgehog family member Sonic hedgehog (Shh) has been shown to regulate cell proliferation and differentiation in the vertebrate retina. Previously, we have reported that Shh secreted by differentiated retinal ganglion cells (RGCs) maintains cell proliferation and negatively regulates RGCs and cone cell genesis in the embryonic mouse retina. Furthermore, we have demonstrated that Math5 mRNA expression is suppressed by Shh signals during RGC genesis using Smoothened (Smo) conditional knockout (cKO) retina. Smo cKO mutants also display increased expression of bHLH genes Hes1 and Ngn2. The purpose of this research is to elucidate the role of Hes1 in Hh signaling in the developing mammalian retina.

Methods: : We have generated retinal specific early deletions of Hes1 and Smo. The conditional gene deletion was performed by crossing mice carrying Smo.flox or Hes.flox allele with Chx10-Cre transgenic mice. In addition, a mouse line with Math5-HA fusion gene knocked-in the Math5 locus was also used to monitor Math5 protein expression in Hes1 and Smo cKO mutant retinas.

Results: : Smo cKO retina shows increased Math5-HA expression in a temporal specific manner. Hedgehog signals are critical in maintaining the cell cycle and inhibiting RGC production in specific progenitor cells. Similarly, Hes1 cKO also results in a decreased progenitor population and increased RGC production. However, Hes1 cKO mutants show earlier and more severe phenotypes than the Smo cKO mutant. We are currently investigating mechanisms underlying changes of Math5 protein and other bHLH genes in Smo and Hes1 cKO mutants.

Conclusions: : These molecular genetic analyses indicate that Shh is an important cell-extrinsic signal that regulates key cell-intrinsic factors during retinal progenitor cell expansion and neuronal fate determination.

Keywords: retinal development • retina • ganglion cells