Embryogenesis is a precisely regulated process that requires tight coordination of events in space but also in time. Despite being exposed to both endogenous and exogenous cues, embryos reliably develop to their final shape within a predictable timeframe. Perturbations have subsequently little effect on embryonic development; i.e. the process of embryogenesis is able to account and to correct for environmental and intra-embryo noise. Despite the observation that development happens in a highly temporally coordinated manner there is little work focusing on how embryos are temporally robust.
Here, we develop procedures to analyze temporal robustness at the whole organism level. Using Drosophila embryos maintained at precise temperatures, we simultaneously image up to 60 embryos from early embryogenesis until larvae hatching. We time the stage of embryogenesis by identifying developmental landmarks. Due to the large embryo number and controlled environment, our setup enables us to study intra- and inter- embryo temporal variations. We analyzed temporal variation at temperatures from 16 to 30°C and found that there exists an optimal developmental temperature (21°C) at which the temporal variation between embryos is minimal. Furthermore, we show that temporal paths are highly correlated at high (>25ºC) and low (<19ºC) temperature (i.e. an embryo that develops quickly at early stages is also fast developing at later stages and vice versa).
In accordance with previous studies, embryos develop faster at higher temperatures, though with increasing temporal variability. Whilst this observation can be explained by the faster kinetics of various processes at increasing temperature, we are now investigating whether active temporal regulators are present and coordinate tissue growth with developmental time. Coupled with microarray analysis, our setup offers the unique opportunity to discover such unknown regulators. We performed miRNA arrays and we found that 22 miRNAs are differentially regulated at varying temperature during embryogenesis. These miRNAs have a broad spectrum of action. Despite being expressed during embryogenesis their absence does not prevent hatching. Hence, they may have a subtler role, possibly ensuring coordination between developmental processes. We are currently using miRNA knockout and miRNA overexpression to further understand their role in controlling developmental time.
Our work represents the first quantitative analysis of temporal variation in embryogenesis and the setup we have developed enables us to explore if – and if so, how – time is effectively regulated during embryogenesis.