Stable, Precise, and Reproducible Patterning of Bicoid and Hunchback Molecules in the Early Drosophila Embryo

Precise patterning of morphogen molecules and their accurate reading out are of key importance in embryonic development. Recent experiments have visualized distributions of proteins in developing embryos and shown that the gradient of concentration of Bicoid morphogen in Drosophila embryos is established rapidly after fertilization and remains stable through syncytial mitoses. This stable Bicoid gradient is read out in a precise way to distribute Hunchback with small fluctuations in each embryo and in a reproducible way, with small embryo-to-embryo fluctuation. The mechanisms of such stable, precise, and reproducible patterning through noisy cellular processes, however, still remain mysterious. To address these issues, here we develop the one- and three-dimensional stochastic models of the early Drosophila embryo. The simulated results show that the fluctuation in expression of the hunchback gene is dominated by the random arrival of Bicoid at the hunchback enhancer. Slow diffusion of Hunchback protein, however, averages out this intense fluctuation, leading to the precise patterning of distribution of Hunchback without loss of sharpness of the boundary of its distribution. The coordinated rates of diffusion and transport of input Bicoid and output Hunchback play decisive roles in suppressing fluctuations arising from the dynamical structure change in embryos and those arising from the random diffusion of molecules, and give rise to the stable, precise, and reproducible patterning of Bicoid and Hunchback distributions

Spatial Bistability Generates hunchback Expression Sharpness in the Drosophila Embryo

During embryonic development, the positional information provided by concentration gradients of maternal factors directs pattern formation by providing spatially dependent cues for gene expression. In the fruit fly, Drosophila melanogaster, a classic example of this is the sharp on–off activation of the hunchback (hb) gene at midembryo, in response to local concentrations of the smooth anterior–posterior Bicoid (Bcd) gradient. The regulatory region for hb contains multiple binding sites for the Bcd protein as well as multiple binding sites for the Hb protein. Some previous studies have suggested that Bcd is sufficient for properly sharpened Hb expression, yet other evidence suggests a need for additional regulation. We experimentally quantified the dynamics of hb gene expression in flies that were wild-type, were mutant for hb self-regulation or Bcd binding, or contained an artificial promoter construct consisting of six Bcd and two Hb sites. In addition to these experiments, we developed a reaction–diffusion model of hb transcription, with Bcd cooperative binding and hb self-regulation, and used Zero Eigenvalue Analysis to look for multiple stationary states in the reaction network. Our model reproduces the hb developmental dynamics and correctly predicts the mutant patterns. Analysis of our model indicates that the Hb sharpness can be produced by spatial bistability, in which hb self-regulation produces two stable levels of expression. In the absence of self-regulation, the bistable behavior vanishes and Hb sharpness is disrupted. Bcd cooperative binding affects the position where bistability occurs but is not itself sufficient for a sharp Hb pattern. Our results show that the control of Hb sharpness and positioning, by hb self-regulation and Bcd cooperativity, respectively, are separate processes that can be altered independently. Our model, which matches the changes in Hb position and sharpness observed in different experiments, provides a theoretical framework for understanding the data and in particular indicates that spatial bistability can play a central role in threshold-dependent reading mechanisms of positional information

Measuring variability

Gene expression is a noisy stochastic process, since it involves at its core interactions between single molecules: a polymerase and a binding site. However, many biological processes directly dependent upon gene expression are reliable. Prominent among them is morphogenesis: how are body parts so consistently generated and proportioned? In the early embryo, gradients of certain proteins called morphogens affect the pattern of cell differentiation and embryonic development. The variability in morphogen patterns and its effect in the proportions of the embryo has been intriguing biologists for a long time, but the limitations, variability and limited reproducibility of immunostaining of fixed embryos does not allow dynamic measurements. New tools now allow precise measurement of the variability of morphogen patterning in living Drosophila embryos, making it possible to probe the underlying mechanisms of development

Morphogen gradient formation and action: Insights from studying Bicoid protein degradation

In a recent publication,1 we identified a novel F-box protein, encoded by fates-shifted (fsd), that plays a role in targeting Bcd for ubiquitination and degradation. Our analysis of mutant Drosophila embryos suggests that Bcd protein degradation is important for proper gradient formation and developmental fate specification. Here we describe further experiments that lead to an estimate of Bcd half-life, <15 min, in embryos during the time of gradient formation. We use our findings to evaluate different models of Bcd gradient formation. With this new estimate, we simulate the Bcd gradient formation process in our own biologically realistic 2-D model. Finally, we discuss the role of Bcd-encoded positional information in controlling the positioning and precision of developmental decisions