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Genetic analysis of protein kinase B (AKT) in Drosophila
1Ontario Cancer Institute, Department of Medical Biophysics, University of Toronto, Canada.
Abstract:
The decision between survival and death is an important aspect of cellular regulation during development and malignancy. Central to this regulation is the process of apoptosis, which is conserved in multicellular organisms [1]. A variety of signalling cascades have been implicated in modulation of apoptosis, including the phosphatidylinositol (Pl) 3-kinase pathway. Activation of Pl 3-kinase is protective, and inhibition of this lipid kinase enhances cell death under several conditions including deregulated expression of c-Myc, neurotrophin withdrawal and anoikis [2-7]. Recently, the protective effects of Pl 3-kinase have been linked to its activation of the pleckstrin homology (PH)-domain-containing protein kinase B (PKB or AKT) [8]. PKB/AKT was identified from an oncogene, v-akt, found in a rodent T-cell lymphoma [9]. To initiate a genetic analysis of PKB, we have isolated and characterized a Drosophila PKB/AKT mutant (termed Dakt1) that exhibits ectopic apoptosis during embryogenesis as judged by induction of membrane blebbing, DNA fragmentation and macrophage infiltration. Apoptosis caused by loss of Dakt function is rescued by caspase suppression but is distinct from the previously described reaper/grim/hid functions. These data implicate Dakt1 as a cell survival gene in Drosophila, consistent with cell protection studies in mammals.
Insights
The phosphatidylinositol 3-kinase (PI3K) pathway promotes cell survival. A Drosophila mutant lacking the PKB/AKT gene, Dakt1, shows increased apoptosis, indicating Dakt1 is crucial for cell survival.
Area of Science:
- Cellular Biology
- Developmental Biology
- Genetics
Background:
- Apoptosis is a fundamental cellular process regulating survival and death during development and disease.
- The phosphatidylinositol 3-kinase (PI3K) pathway plays a critical role in cell survival, with its activation being protective against cell death.
- Protein kinase B (PKB or AKT), activated by PI3K, is a key mediator of these survival signals, with its discovery linked to an oncogene.
Purpose of the Study:
- To genetically analyze the function of PKB/AKT in a model organism.
- To investigate the role of Dakt1, a Drosophila homolog of PKB/AKT, in cellular regulation.
Main Methods:
- Isolation and characterization of a Drosophila PKB/AKT mutant (Dakt1).
- Assessment of apoptosis through indicators like membrane blebbing, DNA fragmentation, and macrophage infiltration during embryogenesis.
- Genetic analysis involving caspase suppression to evaluate rescue of apoptosis.
Main Results:
- The Dakt1 mutant exhibited ectopic apoptosis during Drosophila embryogenesis.
- Loss of Dakt function led to increased cell death, evidenced by membrane blebbing and DNA fragmentation.
- Apoptosis in Dakt1 mutants was rescued by caspase suppression but was independent of known reaper/grim/hid pathways.
Conclusions:
- Dakt1 functions as a critical cell survival gene in Drosophila.
- These findings in Drosophila are consistent with the known cell-protective roles of PKB/AKT in mammals.
- The study implicates Dakt1 in the conserved regulation of cell survival and apoptosis.