Genetic regulation of cell extrusion in caenorhabditis elegans

Abstract

Programmed elimination of cells occurs during animal development and homeostasis to maintain appropriate cell numbers. One evolutionarily conserved method by which organisms eliminate cells in a programmed manner is by cell-autonomous activation of the caspase-mediated apoptosis pathway, which produces a corpse that is engulfed and degraded by phagocytic cells. Cell elimination can also occur by a different method, called cell extrusion, in which the cell to be eliminated is squeezed out from a layer of cells, such as an epithelium. Cell extrusion is also an evolutionarily conserved form of cell elimination and has been observed in organisms ranging from Drosophila to mammals. It is the primary method of cell elimination in mammalian epithelial tissues, such as the small intestinal epithelium. A specific set of cells is eliminated by cell extrusion by caspase-deficient C. elegans embryos.

Remarkably, these cells show morphological and cytological features of apoptosis in the complete absence of caspases. To identify the genes required for cell extrusion, I performed a genome-scale RNAi screen for worms that express a phenotype arising from defective extrusion. This RNAi screen revealed that genes required for entry into the cell cycle and G1/S-phase transition are required for cell extrusion. From subsequent live-imaging experiments using confocal microscopy, I discovered that cells fated for extrusion earlier entered the cell cycle and arrested in S phase. I found that extruded cells are the much smaller sisters generated by unequal cell divisions. Taken together, my findings indicate that generation from an unequal cell division, entry into the cell cycle, and the S-phase arrest that likely results from these processes are all coupled to the cell extrusion fate, as genetically perturbing any of these processes blocks cell extrusion.

In short, I have identified the genetic factors that lead to the arrested cell-cycle state that drives caspase-independent apoptotic cell extrusion by C. elegans. Studies of mammalian epithelial cells using hydroxyurea (performed in collaboration with the Jody Rosenblatt Laboratory) indicate that S-phase arrest drives cell extrusion from mammalian epithelia. These findings demonstrate that cell extrusion driven by S-phase arrest is evolutionarily conserved and suggest implications for development, physiology and disease.