Jeremy Wickenheiser et al. (2004) West Coast Worm Meeting "An RNAi Screen for CED-4-Dependent Lethal Genes"

Joel H Rothman, Jeremy Wickenheiser

[West Coast Worm Meeting, 2004]

The machinery that activates programmed cell death (PCD) appears to be present in many, if not all cells that do not normally die. It is therefore essential to repress PCD in cells destined to survive. Many of the molecular events that repress PCD in living cells are not well defined and we are performing studies to identify and characterize novel mechanisms that keep PCD inactive in surviving cells. Genetic studies have shown that the egl-1 , ced-3 , and ced-4 genes are necessary for nearly all PCD in C. elegans . We recently reported that the ICD-1 apoptotic suppressor inhibits CED-4-dependent, CED-3-independent PCD (Bloss, et al., Nature 424, 2003), revealing that the pro-apoptotic actions of CED-3 and CED-4 are separable. Derepression of the cell death program in essential cells, as seen in mutants defective for the ced-9 apoptotic suppressor, leads to lethality that is suppressible by mutations in the pro-apoptotic genes ced-3 and ced-4 . In an effort to identify additional PCD suppressors, we have performed an RNAi screen for lethal mutants that are suppressed by a ced-4(-) mutation. From a screen of ~900 genes that exhibit RNAi-induced embryonic or larval lethality, we have identified >40 candidate genes that show suppression of lethality, ranging from mild to >100-fold suppression in a ced-4 mutant. Progress on characterization of these newly identified CED-4-dependent lethal genes will be reported.

Ashish Kumar et al. (2007) International Worm Meeting "Identification of lethal genes dependent on the cell death machinery."

Ashish Kumar, Joel H. Rothman, Jeremy Wickenheiser

[International Worm Meeting, 2007]

Development and maintenance of tissues in animals depends not only on the ability of cells to reproduce, but also on their capacity for self-destruction when they become superfluous, damaged, or otherwise harmful. A tight balance between cell survival and cell death is of fundamental importance for proper development. Genetic analyses have identified an evolutionarily conserved core programmed cell death (PCD) pathway; however, many observations suggest that there is a complex network of genes that ensure that this pathway is activated only at the proper time and position. The pathways by which the proapoptotic factors, such as caspases, are maintained in an inactive state in cells destined to survive are not fully understood. We are performing studies to identify and characterize novel mechanisms that inhibit the apoptotic death of essential cells during development. Activation of the cell death program in essential cells, as observed in mutants defective for the CED-9/bcl-2 cell death inhibitor, leads to lethality that is suppressible by mutations in the pro-apoptotic genes ced-3 and ced-4. In an effort to identify additional cell death inhibitors, we have performed an RNAi screen for lethal mutants that are suppressed by a ced-4(-) mutation. We are investigating genes that when inhibited by RNAi result in a significant decrease in lethality in a ced-4(-) background compared to N2. The identified mutants are being tested further for cell death, proliferation and differentiation defects. From a screen of ~1100 genes that exhibit RNAi-induced embryonic or larval lethality, we have identified ~80 candidate genes that show suppression of lethality, ranging from mild to >70-fold suppression in a ced-4 mutant. Bioinformatics studies suggest that the candidates from the screen fall into distinct genetic and/or functional network clusters. The results of the screen and detailed analysis of candidate genes will be presented.