Single-cell RNA profiling of blastemal senescent cells

Axolotl limb regeneration proceeds through the formation of a blastema, a mound of progenitor cells that accumulate at the end of the amputated stump. These progenitor cells expand and later undergo patterning to regenerate the missing limb, restoring both form and function. A subset of cells within the blastema become senescent, a state of permanent growth arrest. Here, we address the functional relevance of cellular senescence to axolotl limb regeneration, through a combination of gain- and loss-of-function assays. Using transcriptomic analyses on in vitro and in vivo senescent cells, we gain insights into the basis of the senescent phenotype, cell-cycle arrest, and molecular mediators involved in axolotl regeneration at the molecular level. Overall design: Here, we performed gene expression profiling for endogenous blastemal senescent (Rho+) cells at the single-cell level. Senescent cells were isolated on the basis of GalNP-rho labelling from blastemas derived at different stages of regeneration (mid, late, and palette), and analyzed through single-cell RNA-seq.