Reaktionen der Zellzyklusregulation auf Sauerstoffmangel und Kälte in Drosophila
Final Report Abstract
The present conspicuous debate about anthropogenic climate change also increases public awareness of the profound effects of environmental conditions on living entities. Ambient temperature and oxygen levels can fluctuate considerably in certain locations. Such variations have very strong effects on cells. However, the cellular responses to changes in temperature and oxygen concentration, the focus of our research project, are not yet understood very well at the molecular level. To study cellular and molecular responses we have experimented with the fruit fly Drosophila melanogaster. Thereby we were able to exploit the powerful genetic approaches established in this model organism. Moreover, to reduce problem complexity, we have primarily focused on early embryos. In contrast to later developmental stages (larvae, adults), embryos are immotile and hence cannot mount behavioural responses. In addition, specialized organ systems (like the nervous system, fat body, Malpighian tubules, tracheae, i.e., the insect versions of liver, kidney and lungs) are not yet differentiated. Metabolic responses are therefore still uniform and not yet diversified in cell-type specific manners. Finally, at the start of embryogenesis, when maternally provided provisions deposited in the egg during oogenesis are used, not even the transcriptional response levels are available. This latter fact presumably explains our finding that early embryogenesis is the most cold-sensitive stage of the D. melanogaster life cycle. Like most animal species, D. melanogaster belongs to the ectotherms where body temperature conforms to the ambient temperature. Cells in some ectotherms can function over a surprisingly wide range of ambient temperatures even though biological consequences of temperature change are very complex. Diffusion is less affected by temperature than the diverse enzymatic reactions which all have distinct individual temperature profiles. Hence thermal fluctuations pose a formidable challenge to ectotherm organisms. While the heat shock response has been studied extensively, how cells cope with cold has hardly been addressed. Using a unique microscopic set-up for in vivo imaging we have explored low temperature effects on progression through early D. melanogaster embryogenesis. This has led us to the novel finding that nucleocytoplasmic posttranslational modification of proteins with O-linked N-acetyl-D-glucosamine (O-GlcNAc) is closely correlated with ambient temperature not only in D. melanogaster but also in distantly related ectotherms like the nematode Caenorhabditis elegans and the fish Danio rerio. Genetic analyses revealed the importance of high O-GlcNAc levels for successful development at elevated temperatures. As very many cytoplasmic and nuclear proteins in diverse pathways are known to be O-GlcNAc targets, temperature-dependent regulation of this modification might contribute to a coordinate adjustment of diverse cellular processes in response to thermal change. The other project part was focussed on hypoxia effects on Mps1 because this protein had been implicated in the response to oxygen deprivation by our previous work. Mps1 functions in the spindle assembly checkpoint (SAC), a surveillance mechanisms which effectively curbs chromosome missegregation during mitotic divisions and thereby makes an important contribution to maintenance of genetic stability during cell proliferation. We were able to demonstrate that oxygen deprivation induces SAC activation and Mps1 relocalization within minutes in early D. melanogaster embryos. We also observed rapid effects on microtubule organization in both wild-type and Mps1 mutants. Therefore, anoxia effects on the mitotic spindle are up- rather than downstream of SAC activation. Oxygen deprivation impairs the chromosome segregation machinery more rapidly than SAC function. Thus SAC can be activated by spindle damage in response to acute anoxia. Importantly, compromised SAC function has been proposed to contribute to the dramatic chromosome instability (CIN) characteristically observed in most solid tumours in humans. Our results suggest that the hypoxic conditions typical of prevascularized early tumour stages might synergize with a compromised SAC to drive CIN.
Publications
- 2007. Rapid effects of acute anoxia on spindle kinetochore interactions activate the mitotic spindle checkpoint. J Cell Sci. 120:2807-18
Pandey, R., Heeger, S., Lehner, C.F.
