LIN-3/EGF Promotes the Programmed Cell Death of Specific Cells in Caenorhabditis elegans by Transcriptional Activation of the Pro-apoptotic Gene egl-1
Figure 6
lin-3 promotes PCD through transcriptional activation of egl-1 by LIN-1.
(A) The alignment between the C. elegans and C. briggsae genomic sequences near the P3 fragment. *indicates the LIN-1 minimal recognition sequence GGAA/T. (B) GST::LIN-1(DBD) directly binds to the egl-1 promoter in an EMSA assay. Lane 1, no protein; lane 2, 200 ng of GST; lanes 3–5, 10, 50, or 200 ng of GST::LIN-1(DBD); lane 6: 200 ng of GST::LIN-1(DBD)+50× wild-type cold probe; lane 7, 200 ng of GST::LIN-1(DBD)+100× wild-type cold probe; lane 8, 200 ng of GST::LIN-1(DBD)+50× mutant cold probe; lane 9, 200 ng of GST::LIN-1(DBD)+100× mutant cold probe. The bottom arrow indicates non bound probe and the top arrow DNA bound to GST::LIN-1(DBD) and displaying retarded mobility. (C) The percentage of animals with surviving ABpl/rpppapp of the indicated genotypes was scored (n≥30). Ex[egl-1(+)] and Ex[egl-1(mut)] indicate wild-type and mutant egl-1 transgenes, respectively, as described in the text. The surviving ABpl/rpppapp was scored using the sur-5::gfp reporter as described in Figure 1B. Two independent stably transmitted lines were analyzed for each transgene. Alleles used: lin-3(e1417) and egl-1(n1084n3082). *indicates P<0.05 and **P<0.001 when egl-1(lf) and lin-3(lf) were compared to wild-type or the transgenic strains were compared to egl-1(lf); Ex[egl-1(+)] (Fisher's exact test). ns, no significance. (D) Cell corpses in the embryos of the indicated genotypes carrying the wild-type or mutant egl-1 gene were scored at the 1.5-fold stage (n≥30). Each circle represents a single embryo. At least three independent stably transmitted lines were analyzed for each transgene. Alleles used: lin-3(e1417) and egl-1(n1084n3082). **indicates P<0.001 in a two-tailed t-test. ns indicates no statistical difference (p>0.05). (E) Proposed model of how lin-3 promotes PCD (see text for details).