Rif1 prolongs the embryonic S phase at the Drosophila mid-blastula transition
(A) Diagram highlighting the key changes to the cell cycle and to embryonic morphology during early development. The initial prolongation of the cell cycle is due to the increase in the length of S phase. This occurs gradually during cycles 11–13, and then the onset of a late-replication program in S phase 14 extends interphase considerably. The first G2 is introduced in cycle 14, after which cells enter mitosis 14 asynchronously, according to a developmental pattern. (B) Nuclei from an S phase 14 Rif1-GFP embryo stained for GFP (green in the merge) and DNA (DAPI: blue in the merge). Rif1 colocalizes with regions of the interphase nucleus that stain intensely with DAPI. (C) Specific satellites (magenta in the merge), either 1.686 detected live by TALE-light or AATAC detected by DNA FISH colocalized with Rif1-GFP (green in merge) during early cycle 14. (D) Frames from live imaging (accompanied by S1 Movie) showing Rif1-GFP and His2Av-RFP in developing embryos during cell cycles 11–14. Note the initial presence and increasing persistence of nuclear Rif1 foci as interphase lengthens. (E) Magnified images from time-lapse records of Rif1-GFP during S phase 14. Individual nuclear foci of Rif1 disappeared at different times across S phase. (F) Frames from time lapse showing Rif1-GFP in a single nucleus at 20-s intervals during S phase 13 (accompanied by S2 Movie). Arrows indicate 2 specific foci at the time that their fluorescence declined. FISH, fluorescence in situ hybridization; GFP, green fluorescent protein; MBT, mid-blastula transition; His2Av, histone 2A variant; RFP, red fluorescent protein; Rif1, Rap1 interacting factor 1; TALE, transcription activator-like effector.