Fig. 3. Activity of Xath5 as an RGC determinant is affected by modulation of cell cycle activity. (A) Cell cycle exit of RGC precursors is unaffected by Xath5. Xath5 plus GFP or GFP only was lipofected at stage 15, and then BrdU injection started at stage 28 or late stage 32. At stage 41, the ratio of BrdU-positive cells in all lipofected cells or lipofected RGCs was analysed. (B) Cell cycle exit of RGC precursors is affected by p27Xic1 and cyclin E1. p27Xic1 or cyclin E1 was colipofected with Xath5 at stage 15, and then BrdU injection started at early stage 32. At stage 41, the ratio of BrdU-positive lipofected RGCs to all lipofected RGCs was analysed. (C) Activity of Xath5 as a RGC determinant is affected by modulation of cell cycle activity. Xath5 was colipofected with p27Xic1 or cyclin E1 at stage 15, and then their effect on cell fate determination was analyzed. (D) Deleted constructs of p27Xic1 also increase the fraction of cells that become RGCs. (E) Overexpression of cyclin E1 in retinal precursor cells transiently activates the cell cycle. An expression construct of cyclin E1 was lipofected at stage 15 and BrdU injection started at the indicated stages. At stage 42, BrdU incorporation was analyzed. (F-H) Retina lipofected with Xath5 by itself and in combination with cyclin E1. (F) Retina lipofected with GFP. R, RGC; P, photoreceptor cell; H, horizontal cell; A, amacrine cell; B, bipolar cell; M, Müller glial cell. (G) Retina lipofected with Xath5 and GFP. The ratio of RGCs is 50% (12 RGCs out of 24 lipofected cells). (H) Retina lipofected with Xath5, cyclin E1 and GFP. The ratio of RGCs is 22% (11 RGCs out of 50 lipofected cells).