– BioSystems 87 (2007) 191–203 - The cAMP Response Element Binding Protein (CREB) as an integrative HUB selector in metazoans: clues from the Hydra model system Simona CHERA, Kostas KALOULIS and Brigitte GALLIOT° Department of Zoology and Animal Biology, University of Geneva, Sciences III, 30 Quai Ernest Ansermet, CH-1211 Geneve 4, Switzerland. °brigitte.galliot@zoo.unige.ch Summary In eukaryotic cells, a multiplicity of extra-cellular signals can activate a unique signal transduction system that at the nuclear level will turn on a variety of target genes, eliciting thus diverse responses adapted to the initial signal. How distinct signals can converge on a unique signalling pathway that will nevertheless produce signal- specific responses provides a theoretical paradox that can be traced back early in evolution. In bilaterians, the CREB pathway connects diverse extra-cellular signals via cytoplasmic kinases to the CREB transcription factor and the CBP co-activator, regulating according to the context, cell survival, cell proliferation, cell differentiation, pro-apoptosis, long-term memory, hence achieving a « hub » function for cellular and developmental processes. In hydra, the CREB pathway is highly conserved and activated during early head regeneration through RSK- dependent CREB phosphorylation. We show here that the CREB transcription factor and the RSK kinase are co- expressed in all three hydra cell lineages including dividing interstitial stem cells, proliferating nematoblasts, proliferating spermatogonia and spermatocytes, differentiating and mature neurons as well as ectodermal and endodermal myoepithelial cells. In addition, CREB gene expression is specifically up-regulated during early regeneration and early budding. When the CREB function was chemically prevented, the early post-amputation induction of gene as HyBraI was no longer observed and head regeneration was stacked. Thus, in hydra, the CREB pathway appears already involved in multiple tasks, such as reactivation of developmental programs in an adult context, self-renewal of stem cells, proliferation of progenitors and neurogenesis. The hub function played by the CREB pathway established early in animal evolution might have contributed to the formation of an efficient oral pole through the integration of the neurogenic and patterning functions. Keywords : hydra, reactivation of developmental program, genetic control of regeneration, MAPK/RSK/CREB signalling pathway, neurogenesis, patterning processes 1. INTRODUCTION 1.1. The same genetic tools are used in various animal phyla to control development Thirty years ago, a theoretical framework for the genetic control of developmental processes was put forward by Antonio Garcia-Bellido (1975): using the development of the Drosophila wing as a model system, he proposed that, depending on the cell location, a set of selector genes, would be turned on by extra-cellular signals and activate cytodifferentiation genes, also named realisator genes. Similarly, relying on genetic evidences but prior to any molecular characterisation, Ed Lewis proposed that specification of the different segments of the Drosophila embryo was achieved by a segment-specific combination of homeotic gene products that would drive expression of target genes through an evolutionarily-conserved DNA-binding domain Lewis (1978), implying that selector genes were working as transcription factors. Several years later, the cloning of the first Drosophila homeotic genes verified these assumptions: their sequences shared a highly-conserved 180 bp long DNA stretch named homeobox McGinnis et al. (1984), Scott and Weiner (1984), that was shown to encode a DNA-binding domain Shepherd et al. (1984). Moreoever, in the same year, this homeobox was detected as multiple copies in the genome of other metazoans Carrasco et al. (1984), McGinnis et al. (1984). Thereafter, it was demonstrated that selector proteins do act as transcription factors, i.e. in the nucleus, they regulate the transcriptional activity of target genes through specific binding onto short DNA sequences, named regulatory elements or enhancers. Moreover, they interact with various partners, named cofactors, which can increase their selectivity and restrict their activity to specific cell fields or cell stages. The next 15 years showed that signalling pathways are in a limited number, reused many times throughout development and highly conserved among metazoans. Hence, the regulatory elements recognised by transcription factors only respond to a combination of