Hypoxia and Nitric Oxide Induce a Rapid, Reversible Cell Cycle Arrest of the Drosophila Syncytial Divisions

Abstract

Cells can respond to reductions in oxygen (hypoxia) by metabolic adaptations, quiescence or cell death (1). The nuclear division cycles of syncytial stage Drosophila melanogaster embryos reversibly arrest upon hypoxia. We examined this rapid arrest in real time using a fusion of green fluorescent protein and histone 2A. In addition to an interphase arrest, mitosis was specifically blocked in metaphase, much like a checkpoint arrest. Nitric oxide, recently proposed as a hypoxia signal in Drosophila, induced a reversible arrest of the nuclear divisions comparable with that induced by hypoxia. Syncytial stage embryos die during prolonged hypoxia, whereas post-gastrulation embryos (cellularized) survive (2, 3). We examined ATP levels and morphology of syncytial and cellularized embryos arrested by hypoxia, nitric oxide, or cyanide. Upon oxygen deprivation, the ATP levels declined only slightly in cellularized embryos and more substantially in syncytial embryos. Reversal of hypoxia restored ATP levels and relieved the cell cycle and developmental arrests. However, morphological abnormalities suggested that syncytial embryos suffered irreversible disruption of developmental programs. Our results suggest that nitric oxide plays a role in the response of the syncytial embryo to hypoxia but that it is not the sole mediator of these responses.Oxygen is essential for the life of most multi-cellular organisms. Limitations in oxygen have profound physiological and health consequences in humans. Prolonged or severe decreases in physiological oxygen resulting from ischemia are a major contributor to morbidity and mortality in stroke and cardiac infarction (4). Cells in pretumorous growths are often deprived of oxygen as a result of insufficient and inefficient vascularization, and this hypoxia (low oxygen) is postulated to play a role in limiting tumor progression (5). The outcome of hypoxia can differ dramatically. For example, a turtle can survive months of hypoxic conditions, whereas the cells of the human brain begin to die following a few minutes of hypoxia (6). These outcomes presumably depend on the cellular responses to hypoxia, which range from metabolic adaptation to quiescence to apoptosis (1). Among the cellular responses to hypoxia is an arrest of the cell cycle (1, 5). Hypoxia is reported to arrest mammalian cells during G 1 , mid-S phase, and G 2 /M. Analysis of the response to hypoxia in Drosophila has similarly revealed an arrest of the cell cycle in mid-S phase and mitosis (2, 3). Despite the importance of these cellular responses, the signals mediating responses to hypoxia are ill defined, and the factors influencing the type of response are poorly understood. What allows some cells to survive when others die?In this study, we examined the responses to hypoxia during Drosophila embryogenesis. The development of Drosophila from egg to larva is remarkably fast and dynamic. Development begins with 13 rapid mitotic divisions in a common cytoplasm, referred to as the syncytial stage. Next, cell membranes invagina...

Oxygen deprivation causes suspended animation in the zebrafish embryo

“…3). That zebrafish embryos arrest in interphase and not mitosis contrasts with studies of Drosophila, where embryos exposed to oxygen deprivation arrest during interphase, prophase, metaphase, and telophase (11,26,27).…”

mRNA cycles through hypoxia-induced stress granules in live Drosophila embryonic muscles

“…Some of the effects of hypoxia are reversible, for example, hypoxia induces cell cycle arrest in Drosophila embryos but cell cycle activity resumes about 20 min after re-establishing normoxia (DiGregorio et al, 2001;Douglas et al, 2001). However, antibody labeling revealed that after 30 minutes of reoxygenation …”

Regulation of multimers via truncated isoforms: a novel mechanism to control nitric-oxide signaling

“…NO regulates cell proliferation in the imaginal discs of developing larvae (Kuzin et al 1996) and in embryos (Wingrove and O'Farrell 1999), participates in the development of the visual system (Gibbs and Truman 1998;Gibbs 2003), induces vesicle release at the neuromuscular junction of larvae (Wildemann and Bicker 1999), controls epithelial fluid secretion by the Malpigian tubules (Dow et al 1994;Broderick et al 2003), triggers the immune response against bacterial pathogens (Nappi et al 2000), induces arrest of nuclear divisions in early embryo in response to oxygen deprivation (DiGregorio et al 2001), and mediates hypoxia-dependent exploratory behavioral responses in larvae (Wingrove and O'Farrell 1999) and hypoxia-induced stasis in embryos (Teodoro and O'Farrell 2003). This wide range of processes mediated by NO requires tight regulation of its production in response to different stimuli.…”

“…In mammals, these changes include blood-vessel relaxation, immune response, cell cycle control, and neurotransmission. In Drosophila, NO has been implicated in visual-system development, immunity, behavior, response to hypoxia, osmoregulation, and regulation of cell cycle progression during development (Dow et al 1994;Kuzin et al 1996Kuzin et al , 2000Gibbs and Truman 1998;Wingrove and O'Farrell 1999;Nappi et al 2000;DiGregorio et al 2001;Teodoro and O'Farrell 2003).…”