Figure 8

Effect of NM23-X4 on cell cycle regulation. (A) BrdU assay in Xenopus retina. The indicated constructs were co-lipofected with GFP at stage 15. BrdU was injected into embryos from stage 30 to 41 and ratios of BrdU positive cells within GFP positive lipofected cells in the inner nuclear layer (INL) and ganglion cell layer (GCL) were determined as described in the Materials and methods. NM23-X4 does not influence proliferation, while shX4-A and -B reduce the ratio of BrdU positive cells. Co-overexpression of NM23-X4 with p27Xic1 inhibits p27Xic1 mediated cell cycle arrest. (B, C) NM23 can inhibit p27Xic1-mediated cell cycle arrest in early embryos. After mRNA injection into a two-cell stage blastomere, the blastomere size of the injected side was compared with the uninjected side at stage 7. The effect was evaluated and the data are presented in the graph shown in (C). (D) A proposed model for NM23-X4 function in retinal cell fate determination. Both NM23-X4 and p27Xic1 are expressed in the CMZ. NM23-X4 expression starts first and then overlaps with p27Xic1 expression. p27Xic1 expression gradually increases in the CMZ [13]. Gliogenic activity is also gradually activated in the ciliary marginal zone. NM23-X4 inhibits the activity of p27Xic1-mediated gliogenesis at the peripheral side of the p27Xic1-expression domain. This results in proper temporal and spatial regulation of gliogenesis. Single, double and triple asterisks correspond to P ≤ 0.05, 0.01, and 0.001, respectively; error bars indicate standard error of the mean.