The tumour suppressor Lethal (2) giant discs is required for the function of the ESCRT-III component Shrub/CHMP4.

Recent work indicates that defects in late phases of the endosomal pathway caused by loss of function of the tumour suppressor gene lethal (2) giant discs (lgd) or the function of the ESCRT complexes I-III result in the ligand-independent activation of the Notch pathway in all imaginal disc cells in Drosophila melanogaster. lgd encodes a member of an uncharacterised protein family, whose members contain one C2 domain and four repeats of the DM14 domain. The function of the DM14 domain is unknown. We here report a detailed structure-function analysis of Lgd protein, which reveals that the DM14 domains are essential for the function of Lgd and act in a redundant manner. Moreover, our analysis indicates that the DM14 domain provides the specific function, whereas the C2 domain is required for the subcellular location of Lgd. We found that Lgd interacts directly with the ESCRT-III subunit Shrub through the DM14 domains. The interaction is required for the function of Shrub, indicating that Lgd contributes to the function of the ESCRT-III complex. Furthermore, our genetic studies indicate that the activation of Notch in ESCRT and lgd mutant cells occurs in a different manner and that the activity of Shrub and other ESCRT components are required for the activation of Notch in lgd mutant cells.

Pan-caspase inhibition suppresses polyethylene particle-induced osteolysis.

Particle-induced osteolysis is a major cause of aseptic loosening after total joint replacement. Earlier studies demonstrated apoptotic macrophages, giant cells, fibroblasts and T-lymphocytes in capsules and interface membranes of patients with aseptic hip implant loosening. The aim of the current study was to determine in a murine calvarial model of wear particle-induced osteolysis whether inhibition of apoptosis using the pan-caspase inhibitor BOC-D-FMK reduces aseptic loosening. Healthy 12-week-old male C57BL/6J mice were treated with UHMWPE particles and received a daily peritoneal injection of BOK-D-FMK, respectively only buffer at a dose of 3 mg/kg of body weight for 12 days until sacrifice. Bone resorption was measured by histomorphometry, micro CT (computed tomography) and TRAP-5b serum analysis. Apoptosis was measured using caspase-3 cleaved staining. The results demonstrated that UHMWPE particles induced stronger apoptotic reactions in macrophages and osteoblasts and increased bone resorption in non-specifically treated mice, whereas peritoneal application of BOC-D-FMK significantly counteracted these adverse particle-related effects. We think that in particle-induced osteolysis apoptosis is pathologically increased, and that failure to reduce the quantity of apoptotic bodies leads to an up-regulation of proinflammatory cytokines, which may be responsible for the induction of osteolysis. We showed for the first time in vivo that a reduction in apoptosis leads to a significant reduction in particle-induced osteolysis. Clinically, the apoptotic cascade could become an interesting novel therapeutic target to modulate particle-induced osteolysis.