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Biochemical and genetic interactions between Drosophila caspases and the proapoptotic genes rpr, hid, and grim
1Departments of Biology and Brain and Cognitive Sciences, Howard Hughes Medical Institute, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Abstract:
In Drosophila melanogaster, the induction of apoptosis requires three closely linked genes, reaper (rpr), head involution defective (hid), and grim. The products of these genes induce apoptosis by activating a caspase pathway. Two very similar Drosophila caspases, DCP-1 and drICE, have been previously identified. We now show that DCP-1 has a substrate specificity that is remarkably similar to those of human caspase 3 and Caenorhabditis elegans CED-3, suggesting that DCP-1 is a death effector caspase. drICE and DCP-1 have similar yet different enzymatic specificities. Although expression of either in cultured cells induces apoptosis, neither protein was able to induce DNA fragmentation in Drosophila SL2 cells. Ectopic expression of a truncated form of dcp-1 (DeltaN-dcp-1) in the developing Drosophila retina under an eye-specific promoter resulted in a small and rough eye phenotype, whereas expression of the full-length dcp-1 (fl-dcp-1) had little effect. On the other hand, expression of either full-length drICE (fl-drICE) or truncated drICE (DeltaN-drICE) in the retina showed no obvious eye phenotype. Although active DCP-1 protein cleaves full-length DCP-1 and full-length drICE in vitro, GMR-DeltaN-dcp-1 did not enhance the eye phenotype of GMR-fl-dcp-1 or GMR-fl-drICE flies. Significantly, GMR-rpr and GMR-grim, but not GMR-hid, dramatically enhanced the eye phenotype of GMR-fl-dcp-1 flies. These results indicate that Reaper and Grim, but not HID, can activate DCP-1 in vivo.
Insights
Reaper (rpr) and Grim proteins activate the Drosophila caspase DCP-1 in vivo, a key step in programmed cell death. This activation is crucial for inducing apoptosis and DNA fragmentation in flies.
Area of Science:
- * Molecular Biology
- * Developmental Biology
- * Genetics
Background:
- * Apoptosis induction in Drosophila melanogaster relies on three genes: reaper (rpr), head involution defective (hid), and grim.
- * These genes activate apoptosis by triggering a caspase pathway, with DCP-1 and drICE being identified Drosophila caspases.
- * DCP-1 exhibits substrate specificity similar to human caspase 3 and C. elegans CED-3, indicating its role as a death effector caspase.
Purpose of the Study:
- * To investigate the in vivo activation of Drosophila caspases DCP-1 and drICE by apoptosis-inducing genes.
- * To determine the functional roles of DCP-1 and drICE in Drosophila development, specifically in the retina.
- * To elucidate the specific roles of reaper, hid, and grim in activating DCP-1.
Main Methods:
- * Ectopic expression of full-length and truncated forms of dcp-1 and drICE in the developing Drosophila retina using an eye-specific promoter.
- * In vitro cleavage assays to assess the interaction between active DCP-1 and full-length DCP-1/drICE.
- * Co-expression of apoptosis-inducing genes (rpr, hid, grim) with dcp-1 to evaluate in vivo activation and phenotypic enhancement.
Main Results:
- * Ectopic expression of truncated dcp-1 (DeltaN-dcp-1) caused a small and rough eye phenotype, while full-length dcp-1 had minimal effect.
- * Expression of full-length or truncated drICE did not result in obvious eye phenotypes.
- * Active DCP-1 cleaved DCP-1 and drICE in vitro, but DeltaN-dcp-1 did not enhance phenotypes of DCP-1 or drICE expression.
- * Co-expression of reaper and grim, but not hid, with dcp-1 significantly enhanced the eye phenotype, indicating in vivo activation of DCP-1.
Conclusions:
- * Reaper and Grim are capable of activating DCP-1 in vivo, playing a critical role in initiating apoptosis.
- * While DCP-1 and drICE share enzymatic specificities, their roles and regulation in vivo differ.
- * The findings highlight the specific roles of Reaper and Grim as direct activators of the death effector caspase DCP-1 in Drosophila.