Ecdysone signaling induces two phases of cell cycle exit inDrosophilacells

Abstract

During development, cell proliferation and differentiation must be tightly coordinated to ensure proper tissue morphogenesis. Because steroid hormones are central regulators of developmental timing, understanding the links between steroid hormone signaling and cell proliferation is crucial to understanding the molecular basis of morphogenesis. Here we examined the mechanism by which the steroid hormone ecdysone regulates the cell cycle in Drosophila. We find that a cell cycle arrest induced by ecdysone in Drosophila cell culture is analogous to a G2 cell cycle arrest observed in the early pupa wing. We show that in the wing, ecdysone signaling at the larva-to-puparium transition induces Broad which in turn represses the cdc25c phosphatase String. The repression of String generates a temporary G2 arrest that synchronizes the cell cycle in the wing epithelium during early pupa wing elongation and flattening. As ecdysone levels decline after the larva-to-puparium pulse during early metamorphosis, Broad expression plummets, allowing String to become re-activated, which promotes rapid G2/M progression and a subsequent synchronized final cell cycle in the wing. In this manner, pulses of ecdysone can both synchronize the final cell cycle and promote the coordinated acquisition of terminal differentiation characteristics in the wing.

Hormone-dependent control of developmental timing through regulation of chromatin accessibility

“…We reasoned that ecdysone signaling may be involved, since it controls developmental transitions in insects (Thummel 2001;King-Jones and Thummel 2005), and our previous work suggested that an extrinsic signal may coordinate temporal changes in chromatin accessibility between the appendages (McKay and Lieb 2013). We performed RNA-seq at six time points in pupal wings (Guo et al 2016). Consistent with the Ashburner model of ecdysone signaling ( Fig.…”

“…During the next 2 d of pupal development (Fig. 1B, C), cell fates continue to be more finely determined, while the wing undergoes a final round of cell division (Guo et al 2016). This time is also characterized by dramatic morphological changes at both the tissue and cellular levels: Changes in cell shape drive eversion of the wing pouch, and changes in cell adhesion allow the apposed dorsal and ventral surfaces of the wing epithelium to form the upper and lower layers of the wing blade.…”

“…1B, "24 h") and 44 h (Fig. 1C, "44 h") after puparium formation (Guo et al 2016). Pairwise comparisons between successive time points revealed thousands of genes both increasing and decreasing between each time point (edgeR false discovery rate [FDR] <0.05; fold change greater than or equal to twofold for expressed genes) ( Fig.…”

“…Last, we examined two candidate enhancers from the broad (br) locus, which encodes a family of transcription factors active in third instar and early prepupal tissues, including the wing (Kiss et al 1988;Guo et al 2016). Using our FAIRE-seq data, we identified open chromatin sites at the br locus that are accessible in L3 wing discs but subsequently decrease in accessibility by 24 h and 44 h ( Fig.…”

“…RNA was prepared as described previously (Guo et al 2016), and the KAPA stranded mRNA-seq kit was used for library construction. FAIRE-seq was performed as described previously (McKay and Lieb 2013), and the Rubicon Thruplex DNA-seq kit was used for library construction.…”