A <em>Drosophila</em> Model to Study Wound-induced Polyploidization

Abstract

Polyploidy is a frequent phenomenon whose impact on organismal health and disease is still poorly understood. A cell is defined as polyploid if it contains more than the diploid copy of its chromosomes, which is a result of endoreplication or cell fusion. In tissue repair, wound-induced polyploidization (WIP) has been found to be a conserved healing strategy from fruit flies to vertebrates. WIP has several advantages over cell proliferation, including resistance to oncogenic growth and genotoxic stress. The challenge has been to identify why polyploid cells arise and how these unique cells function. Provided is a detailed protocol to study WIP in the adult fruit fly epithelium where polyploid cells are generated within 2 days after a puncture wound. Taking advantage of D. melanogaster's extensive genetic tool kit, the genes required to initiate and regulate WIP, including Myc, have begun to be identified. Continued studies using this method can reveal how other genetic and physiological variables including sex, diet, and age regulate and influence WIP's function. 1,2. Postembryonic wound healing in larvae, pupae, and adult fruit flies results in extracellular matrix remodeling, melanin scar formation, and epithelial cell growth 3,4,5,6. The epithelial cells increase in size by cell fusion and the endocycle, an incomplete cell cycle that bypasses mitosis 3,4,7,8. As result, cell loss is compensated by polyploid cell growth instead of cell division. The adult fly hindgut, midgut, and follicular epithelium also rely on polyploid cell growth to compensate for cell loss after tissue damage 9,10,11 .Polyploidy is a well-known aspect of organismal development in plants and insects, but in the last few years it has become more apparent that polyploidy is a conserved tissue repair strategy in vertebrates 12 . The zebrafish, which has the capacity to regenerate its heart, relies on polyploid cell growth to heal damaged epicardium 13 . Polyploidy also contributes to mammalian liver regeneration and kidney tubule epithelium repair after acute injury 14,15 . In these examples, polyploid cells are generated by endoreplication via either endocycle or endomitosis, which results in a binucleated cell due to a block in cytokinesis 12 . The enigma is why polyploid cells arise during wound repair and how polyploidy affects tissue function. Recent studies have provided new insight into the question of whether polyploidy offers a healing advantage or disadvantage. In zebrafish epicardium, polyploidy enhanced the speed of wound healing

Epithelial mechanics are maintained by inhibiting cell fusion with age in Drosophila

“…Newly eclosed flies were collected and aged (males and females together), for the time as denoted in figure legend. Female flies (unless specified as male) were dissected as previously described (Bailey et al, 2020). Briefly, abdomens were fixed in 4% paraformaldehyde, permeabilized in 1× PBS with 0.3% Triton X-100 and 0.3% BSA, then stained overnight at 4°C using primary antibodies.…”

“…Epithelial ploidy was calculated according to Bailey et al (2020). Samples from young and old flies were dissected and stained as described above, then imaged with the same exposure time.…”

“…Adult flies were aged for either 7-9 days or were 40 days old and dissected as described previously (Bailey et al, 2020). Freshly dissected tissue was mounted in halocarbon oil 27 (Sigma).…”