Rif1 prolongs the embryonic S phase at the Drosophila mid-blastula transition
(A) Still images from time-lapse imaging of Rif1-GFP and RFP-HP1a during cycle 14 (early S phase through mitosis). During S phase, the number of Rif1 foci decline as the number of HP1a foci increase. G2 nuclei lack Rif1 foci but retain strong HP1a foci (74 min). During the asynchronous mitosis 14 (74/90/96 min), both proteins are lost but are rapidly recruited to late anaphase chromosomes (90/96 min). (B) Still images from time-lapse imaging of GFP-HP1a protein injected into WT embryos (above) and rif1 embryos (below) during S phase 14. HP1a recruitment to the heterochromatin proceeded similarly in control and mutant embryos. (C) Still images from time-lapse imaging of the replication of the 359-satellite progressing from G2 of cycle 14 until completion of its replication in cycle 15. Note that the TALE-light signal is not immediately visible following mitosis (4:00 min frame). We previously described how HP1a binds to the 359 bp repeat following its replication during S phase 14 and subsequently delays its replication in S phase 15. In rif1 embryos, the 359 repeat also replicated late in S phase 15 (in frame 30:00, yellow arrow indicates where the 359 TALE signal overlapped with PCNA). GFP, green fluorescent protein; HP1a, heterochromatin protein 1a; PCNA, proliferating cell nuclear antigen; RFP, red fluorescent protein; Rif1, Rap1 interacting factor 1; TALE, transcription activator-like effector; WT, wild-type.