TI - Results ABL-1 Inhibits the Engulfment of Apoptotic Cell Corpses . AB - To test whether abl-1 has a role in engulfment , we counted the number of unengulfed apoptotic cell corpses in the heads of first larval stage ( L1 ) animals harboring mutations in abl-1 and engulfment pathway genes . The number of unengulfed corpses varies with the strength of the engulfment defect and defines a quantitative assay of engulfment defects [35] . We used two presumptive null alleles of abl-1 in this study , n1963 and ok171 .n1963 is a G-to-A transition at the splice acceptor of exon 10 ( bp 16967 of the M79 cosmid sequence ) , resulting in removal of most of the kinase domain and the change of a conserved arginine to serine .ok171 is a deletion allele that removes the entire kinase domain , most of the SH2 domain , and results in a frameshift and an opal stop codon 52 bp later [32] . abl-1 mutation alone had no obvious effect on engulfment . However , mutation of abl-1 decreased the number of unengulfed corpses in the heads of ced-1 , ced-6 and ced-7 mutants ( alleles e1735 , n2095 and n1892 , respectively , all of which are nulls ) ( Table 1 ) . dyn-1 mutants die as embryos and were not tested . ABL-1 function did not depend on the presence of functional CED-1 , CED-6 or CED-7 and therefore ABL-1 acts independently or downstream of the CED-1 pathway . These data are consistent with a role for ABL-1 in the negative regulation of apoptotic cell engulfment . Experiments that address alternative explanations for the affect of ABL-1 on engulfment are presented in the next section . We tested for interactions between abl-1 and the genes of the CED-10 Rac pathway .abl-1 mutation did not modify the engulfment defects of either ced-5 ( n1812 ) or ced-12 ( n3261 ) null mutants ( Table 1 ) . abl-1 mutation also did not modify the engulfment defect of ced-2 ( n1994 ) ; mig-2 ( gm38 mu133 ) null double mutants , in which both known inputs into the CED-10 Rac pathway are absent ( Figure 1 ) . However , abl-1 mutation did partially suppress the engulfment defect caused by ced-2 ( n1994 ) alone ( the number of cell corpses decreased from 220 to 157 , p lt 00001 ) and by the partial loss-of-function allele ced-2 ( e1752 ) alone ( a decrease from 189 to 132 , p lt 00001 ) . mig-2 null mutations do not cause engulfment defects on their own so they were tested in combination with a CED-1 pathway mutant ( see below ) . Animals completely lacking ced-10 die as embryos and were not tested , but the engulfment defect caused by a partial loss-of-function allele , ced-10 ( n1993 ) , was suppressed by abl-1 ( lf ) ( the number of cell corpses decreased from 20 to 87 , p lt 00001 ) . We also tested whether the engulfment defect of a ced-1 (e1735) ; ced-2 ( n1994 ) double mutant could be suppressed . A mutation in abl-1 did not modify the engulfment defect of the double mutant , even though each single mutant was suppressed . This result is consistent with the possibility that ABL-1 acts upstream of both the CED-10 Rac and the CED-1 pathways . Alternatively , ABL-1 might suppress a pathway parallel to these two pathways , but its suppression might be too weak to modify an engulfment defect as severe as that of the ced-1 (e1735) ; ced-2 ( n1994 ) double mutant . We present data below that support the latter model . Because mig-2 and unc-73 mutations enhance the engulfment defects of other engulfment gene mutations but do not cause defects on their own [23] , we tested whether ABL-1 acts through the MIG-2 branch of the CED-10 Rac pathway by testing whether an abl-1 mutation could suppress the mig-2 enhancement of ced-1 ( n2091 ) . We observed fewer apoptotic cell corpses in the heads of ced-1 ( n2091 ) ; mig-2 ( gm38 mu133 ) animals when an abl-1 mutation was present ( Table 1 ) , demonstrating that ABL-1 can act in the absence of MIG-2 function . Therefore , ABL-1 does not act solely through either the CED-2 branch or the MIG-2 branch of the CED-10 Rac engulfment pathway . In summary , abl-1 ( lf ) suppressed partial but not complete loss of the CED-10 Rac pathway . The inability of abl-1 ( lf ) to suppress the engulfment defects of the CED-10 Rac pathway when this pathway was completely nonfunctional ( ie , ced-5 null , ced-12 null , or ced-2 ; mig-2 double null mutants , Figure 1 ) suggests that the CED-10 Rac pathway genes do not function by blocking the action of ABL-1 . Instead , ABL-1 might inhibit the CED-10 Rac pathway or ABL-1 might signal in parallel to the CED-10 Rac pathway through another group of effectors that require CED-10 Rac pathway function to accomplish apoptotic cell engulfment . Notably , the observation that abl-1 ( lf ) suppressed the engulfment defect of mutants that completely lack CED-2 function indicates that the effect of ABL-1 on engulfment is atleast partially independent of CED-2 , i.e. , ABL-1 does not act only by inhibiting CED-2 . Therefore , C.elegans ABL-1 can act to inhibit CED-2 CrkII -dependent pathways via a mechanism distinct from the known mechanism in mammals , in which Abl PHOSphorylates the CED-2 homolog CrkII .