Figure 7  The E2f3a Isoform Drives the Differentiation Defect in Rb KO SACs
(A) Schematic diagrams of the mouse WT, E2f3a−/−, and the Cre-recombined floxed E2f3 loci (indicated here as E2f3−/− for simplicity). E2f3a−/− mice lack most of E2f3 exon 1a and part of intron 1a (red dotted box). Arrows indicate PCR primers. Genotyping of an E2f3a+/− mouse is shown on the right.
(B) RT-PCR detection of E2f3a and E2f3b mRNA in the retina. The sequences of primers are 1aF (5′-GCCTCTACACCACGCCACAAG-3′), 1bF (5′-CGGAAATGCCCTTACAGC-3′), and 4R (5′-CTCAGTCACTTCTTTGGACAG-3′). WT retina expresses both E2f3a and E2f3b mRNA. As expected, E2f3a−/− retina lacks E2f3a mRNA and still expresses E2f3b mRNA. E2f3−/− retina lacks full-length E2f3a and E2f3b mRNAs, and instead expresses a truncated mRNA lacking exon 3.
(C) Real-time RT-PCR analysis of E2f genes in P8 retinas of the indicated genotypes. Error bars represent SD of measurements from three animals, and asterisks indicate a significant difference between WT and the indicated genotypes (*, p <0.05; **; p <0.01; ANOVA and Tukey HSD test).
(D) Rescue of Rb KO SACs by E2f3a deletion. Horizontal retinal sections of the indicated genotypes and ages were stained for nuclei (DAPI, blue), M-phase (PH3, green), and the SAC marker Slc18a3 (red). E2f3a deletion does not suppress ectopic division, but rescues the SAC defect. Scale bars are 50 μm.
M, molecular size marker.