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The cnidarian freshwater polyp Hydra sp. exhibits an unparalleled regeneration capacity in the animal kingdom. Using an integrative transcriptomic and stable isotope labeling by amino acids in cell culture proteomic/phosphoproteomic approach, we studied stem cell-based regeneration in Hydra polyps. As major contributors to head regeneration, we identified diverse signaling pathways adopted for the regeneration response as well as enriched novel genes. Our global analysis reveals two distinct molecular cascades: an early injury response and a subsequent, signaling driven patterning of the regenerating tissue. A key factor of the initial injury response is a general stabilization of proteins and a net upregulation of transcripts, which is followed by a subsequent activation cascade of signaling molecules including Wnts and transforming growth factor (TGF) beta-related factors. We observed moderate overlap between the factors contributing to proteomic and transcriptomic responses suggesting a decoupled regulation between the transcriptional and translational levels. Our data also indicate that interstitial stem cells and their derivatives (e.g., neurons) have no major role in Hydra head regeneration. Remarkably, we found an enrichment of evolutionarily more recent genes in the early regeneration response, whereas conserved genes are more enriched in the late phase. In addition, genes specific to the early injury response were enriched in transposon insertions. Genetic dynamicity and taxon-specific factors might therefore play a hitherto underestimated role in Hydra regeneration., (© The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Cnidarians are simple metazoans with only two body layers and a primitive nervous system. They are famous for their nearly indefinite regeneration capacity. Recent work has identified most of the Wnt subfamilies and Wnt antagonists known from vertebrates in this basal animal model. Wnt signaling and BMP signaling have been shown to act in Hydra pattern formation and regeneration. Because recent genomic work in Hydra and Nematostella revealed many genes for vertebrate signaling pathways and transcription factors to be present in this more than 500 Myr-year-old phylum, future work will focus on the function and expression of these genes in Hydra pattern formation and regeneration. This chapter presents an in situ hybridization protocol, which is largely based on a lab protocol of the Bode lab that has proven to be extremely useful in the characterization of many developmental genes from Hydra.