eye disc & mitotic cell cycle
The high levels of apoptosis (detected by Caspase-3 staining) observed in Rbf15aΔ;Stam19 double homozygous somatic clones in third instar larval eye disc is completely abolished when the clones are created in E2f1[i2]/E2f1[rM729] mutant background.
An E2f1rM729 mutant background dominantly enhances the Scer\GAL4GMR.PF>lin-52GD12885 eye phenotype. This is accompanied by a corresponding increase in hid protein and elevated apoptosis in the eye-antennal disc.
The prevailing cell death phenotype observed in Rbf120a Tsc1f01910 double mutant clones at the morphogenetic furrow is no longer present in Rbf120a E2frM729 Tsc1f01910 triple mutant cells, indicating that the increased level of ectopic cell death observed in Rbf120a Tsc1f01910 double mutant cells is E2f-dependent.
Each of l(3)mbt6a14, l(3)mbt4d30, l(3)mbt3d23, l(3)mbt5d8, l(3)mbt4d16, l(3)mbt5d6, l(3)mbt6a31, l(3)mbt3d33, Doa6d2, Doa6a21, Doa5d7, gfzf6a27, B525d21, B523d16, B523d24, CG311337d21, CG31133f07858, bel7d19, Su(E1)-A7d13, Su(E1)-A6a39, Su(E1)-B4d26, Su(E1)-B6a11, Su(E1)-C1d24, Su(E1)-C3d28, Su(E1)-D6a47, Su(E1)-E3d25, Su(E1)-F6b9, Su(E1)-G6a3, Su(E1)-H6a50, Su(E1)-I6d9, Su(E1)-J6a8, Su(E1)-K6b12, Su(E1)-L6b19, Su(E1)-M6b31, Su(E1)-N4d22 and Su(E1)-O6a19 can suppress the proliferation block of E2frM729 clones, allowing recovery of double mutant clones in the eye.
The second mitotic wave is weakly rescued in CG311337d21 E2frM729 and gfzf6a27 E2frM729 double mutant clones in the eye disc, with the double mutant cells entering the second mitotic wave several columns more posterior than in wild-type cells. In addition BrdU incorporation persists several columns more posterior in the double mutant cells than in the adjacent wild-type cells.
The second mitotic wave is weakly rescued in Doa6d2 E2frM729 double mutant clones in the eye disc, with double mutant cells only being sporadically labelled with BrdU in the second mitotic wave. Those double mutant cells that do incorporate BrdU are always found to be significantly more than adjacent wild-type cells that incorporate BrdU.
In wild-type animals, exposure to 40Gy of γ rays induces a distinct pattern of apoptosis in the wing disc, focussed on the the wing pouch, but excluding the DV boundary region and prospective vein. In E2frM729/E2f91; E2f276Q1/E2f2g5 animals, the same irradiation treatment induces a different pattern of apoptosis a smaller number of apoptotic cells are clustered in the center of the wing pouch, many in the D-V boundary region or prospective veins.
The addition of E2f276Q1/Df(2L)G5.1 to E2f91/E2frM729 flies almost completely suppresses the E2f91/E2frM729 phenotypes - mutants develop normally without any significant delay in larval growth, reach pupal stage and finally die as mid- or late pupae. The pattern of DNA synthesis in eye discs is largely normal. The addition of Dpa2/Df(2R)vg-B to E2f91/E2frM729 flies almost completely suppresses the E2f91/E2frM729 phenotypes - mutants develop normally without any significant delay in larval growth.
Approximately 50% of the expected number of Rbf14/Y ; E2fi2/E2frM729 flies survive to the adult stage, and the development of these flies is not significantly delayed compared to sibling flies. The surviving flies have normal eyes, normal macrochaetae on the notum and normal wings and legs. The larval lethality of E2f91/E2frM729 animals can be suppressed by Rbf120a/Rbf14; the larvae develop into large late third instar larvae and many can pupate. Some Rbf120a/Rbf14 ; E2f91/E2frM729 animals can develop into pharate adults with adult eyes, legs and wing structures. The development of these larvae is significantly retarded compared to wild-type larvae, with the earliest pupae observed at around day 11 after egg laying. The normal pattern of DNA replication in the optic lobe region is restored in E2f91/E2frM729 larvae that are also carrying Rbf120a/Rbf14. In Rbf120a/Rbf14 ; E2f91/E2frM729 eye discs the overall pattern of DNA replication is normal. No significant cell death is seen in these eye discs. In late third instar discs there are about 25 rows of developing photoreceptor cells (as in wild-type discs) but there are much fewer cells anterior to the morphogenetic furrow than in wild type.
The early larval lethality of E2f1rM729/Df(3R)Exel6186 transheterozygotes is partially rescues by Scer\GAL4Ubi.PU-driven expression of either E2f1UAS.a.Tag:FLAG,Tag:MYC or E2f1UAS.b.Tag:FLAG,Tag:MYC (giving rise to few, severely developmentally delayed third instar larvae and pupae, but no pharate adults), or of both transgenes (giving rise to the expected numbers of mildly developmentally delayed third instar larvae, pupae and to pharate adults, but to no adults).