The Axolotl as a system to define the function and evolution of reprogramming activities

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Non-mammalian model organisms have been critical in mechanistically dissecting biological processes and identifying the essential, conserved function of genes. It was unknown whether non-mammalian OCT4 and SOX2 orthologs can reprogram cells to pluripotency. The Axolotl is an ideal non-mammalian model to analyze the conservation of OCT4/SOX2 driven reprogramming. AxOCT4 can sustain mouse ES cells. Furthermore the Axolotl is an experimentally accessible model where totipotent germ cells form by embryonic induction. Finally the Axolotl undertakes regeneration that involves reprogramming cells to less committed states.In the last period we showed that AxOCT4 and AxSOX2 can reprogram human fibroblasts to pluripotency. We also showed that OCT4+SOX2+ cells arise in the regenerating spinal cord where cells rebuild functional tissues. Finally, we identified chromatin proteins expressed in regenerating and not wounded tissue likely involved in reprogramming.Here we will further study AxOCT4, AxSOX2 and other factors in reprogramming. First, we will isolate spinal cord OCT4+SOX2+ cells, test their potency, and test the role of OCT4 and our chromatin factors in reprogramming these cells. Second, we will characterize putative ES-like cells cultured from the Axolotl. Finally, we will use transcriptome analysis to compare AxOCT4+SOX2+ cells from atleast three different sources including epiblast and regenerating spinal cord to define the overlapping and non-overlapping gene sets. Comparing this data to the mammalian pluripotency network will refine our understanding of subnetworks required for full pluripotency versus partially reprogrammed states.