Decoding developmental signaling during zebrafish embryogenesis

Eunice Kennedy Shriver National Institute Of Child Health & Human Development

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Abstract

How is signaling molecules distribution regulated? The distribution of signaling molecules within developing tissues helps determine patterns of gene expression. Competing models have been proposed to explain how signaling molecule distributions are established: Signals may diffuse away from producing cells through the extracellular space, move through cells (transcytosis), or be confined to producing cells themselves. We will develop optogenetic tools to probe how factors including extracellular diffusion and transcytosis affect signaling molecule distribution, and use these with in vivo methods including FRAP and FDAP to directly measure signaling molecule mobility and stability. This will help determine how signaling molecule distribution is regulated during zebrafish embryogenesis. How is information encoded in signaling gradients? Gradients of signaling molecules are found in developing tissues from the fly wing precursor to the mammalian neural tube. The classic morphogen model proposes that the precise graded distribution of signaling is important because genes are activated by different signaling levels. Alternatively, a simple signaling asymmetry may suffice to pattern tissues in some contexts. The relatively subtle signaling perturbations required to distinguish between these models can be difficult to achieve in vivo. We will use our optogenetic approaches together with digital micromirror devices to introduce novel signaling distributions in zebrafish embryos and assess patterning consequences. This will determine the spatiotemporal signaling requirements for normal tissue patterning during early zebrafish development. How are signaling levels, dynamics, and combinations interpreted in the embryo? Cells in the developing embryo experience distinct signaling dynamics, levels, and combinations that control differentiation into the diverse cell fates needed in healthy adults. How do different genes respond to these inputs? We seek to determine the input/output relationship between signaling and gene expression during early vertebrate embryogenesis. To achieve this, we are developing orthogonal optogenetic tools to manipulate signaling levels, dynamics, and combinations in zebrafish embryos. We will characterize gene responses and investigate the DNA-level mechanisms responsible for differential responses. This will help elucidate which features of signaling encode information, and explain how the diverse gene expression patterns needed to produce healthy adults are robustly generated.