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- Forsberg, Jeremy, et al.
(författare)
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A caspase-2-RFXANK interaction and its implication for MHC class II expression
- 2018
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Ingår i: Cell Death and Disease. - : Springer Science and Business Media LLC. - 2041-4889. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Despite recent achievements implicating caspase-2 in tumor suppression, the enzyme stands out from the apoptotic caspase family as a factor whose function requires further clarification. To specify enzyme characteristics through the definition of interacting proteins in apoptotic or non-apoptotic settings, a yeast 2-hybrid (Y2H) screen was performed using the full-length protein as bait. The current report describes the analysis of a captured prey and putative novel caspase-2 interacting factor, the regulatory factor X-associated ankyrin-containing protein (RFXANK), previously associated with CIITA, the transactivator regulating cell-type specificity and inducibility of MHC class II gene expression. The interaction between caspase-2 and RFXANK was verified by co-immunoprecipitations using both exogenous and endogenous proteins, where the latter approach suggested that binding of the components occurs in the cytoplasm. Cellular co-localization was confirmed by transfection of fluorescently conjugated proteins. Enhanced caspase-2 processing in RFXANK-overexpressing HEK293T cells treated with chemotherapeutic agents further supported Y2H data. Yet, no distinct differences with respect to MHC class II expression were observed in plasma membranes of antigen-presenting cells derived from wild type and caspase-2(-/-) mice. In contrast, increased levels of the total MHC class II protein was evident in protein lysates from caspase-2 RNAi-silenced leukemia cell lines and B-cells isolated from gene-targeted mice. Together, these data identify a novel caspase-2-interacting factor, RFXANK, and indicate a potential non-apoptotic role for the enzyme in the control of MHC class II gene regulation.
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- Ivanova, Elena V.
(författare)
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Caspase activation in human neuroblastoma cells: mechanisms and spatiotemporal aspects
- 2015
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Apoptosis is one of the modes of programmed cell death, in which several members of the caspase family of proteases play the central role. However, activation of apoptotic caspases does not necessarily lead to cell death. Instead, these caspases may mediate, for instance, differentiation or synaptic plasticity, if their activity is restricted in space and time. Such localized caspase activation has been also implicated in the initial stages of neurodegeneration. In order to assess this kind of events at a subcellular level, our research group has previously constructed tau-anchored FRET-based caspase sensors (tAFSs). Here, we demonstrate that localization of tAFSs to the cytoskeleton results in enrichment of the sensors in neuritic processes and enables increased spatiotemporal resolution for live cell imaging of caspase activation, as compared to soluble FRET sensors. This feature is particularly beneficial for investigation of neurodegeneration-related processes.tAFSs were further employed for investigation of caspase activation in neuroblastoma, an extracranial solid pediatric tumor. Tumor necrosis factor-related apoptosis inducing factor (TRAIL) is a promising candidate for cancer treatment due to its ability to selectively trigger apoptosis malignant cells. However, many cancer cells, including neuroblastoma, acquire resistance to TRAIL. Here, we show that in S-type neuroblastoma cell lines, TRAIL resistance is dependent on incomplete activation of apoptotic caspase-3. Sensitization to TRAIL was achieved with protein kinase C (PKC)-inhibiting compounds, suggesting a role for this kinase in blocking the apoptotic response to TRAIL. This effect of PKC could possibly involve stabilization of XIAP, an endogenous caspase inhibitor, as PKC inhibition, in combination with TRAIL treatment, led to downregulation of XIAP.
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- Jangamreddy, Jaganmohan Reddy
(författare)
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Cancer and cancer stem cell targeting agents : A focus on salinomycin and apoptin
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Current cancer treatments involving surgery, radiotherapy, and chemotherapy target the vast majority of cancer cells, but they are only partially effective in eliminating the disease. Failure to eliminate cancer with conventional treatments can lead to recurrence, which usually kills patient. This often occurs when cancer cells develop resistance to cancer drugs or when cancer-initiating cells (cancer stem cells), unaffected by existing treatment procedures, are present. Here, we studied two drugs, salinomycin and apoptin, that exhibit great potential in the future of cancer treatment not only for restricting malignancy, but also in preventing tumor recurrence. Salinomycin is an antibiotic that was used in poultry farming that is now used clinically to target cancer stem cells, and apoptin is a chicken anemia virus-derived protein that is capable of detecting and killing transformed cells. In this study, we delved into the molecular mechanism of salinomycin action leading to cancer cell death. We showed that salinomycin induces autophagy in both cancer and normal primary cells. We further demonstrated that salinomycin promotes mitochondrial fission, thus increasing mitochondrial mass and mitochondria-specific autophagy, mitophagy. Salinomycin-induced cell death was both necrotic and apoptotic as determined by increased release of HMGB1 and caspase-3, -8 and -9 activation. We also found that stress responses of normal and cancer cells to salinomycin differ and this difference is aggravated by starvation conditions. We proposed that a combinational treatment with glucose starvation, or glucose analogues such as 2DG or 2FDG, might enhance the effects of salinomycin on cancer cells while protecting normal cells. We previously reported that apoptin interacts with BCRABL1, a protein that is expressed in patients with chronic myeloid leukemia (CML). We located a minimal region on the apoptin protein that triggers inhibition of downstream BCR-ABL1 signaling effects. This deca-peptide region was tested on patient samples and was shown to effectively kill cancer cells derived from patients, similar to the drug Imatinib. We further show that the apoptin decapeptide is cytotoxic to Imatinib-resistant patient-derived cancer cells. Thus, we identified a novel therapeutic targeting agent that can not only overcome drug resistance, but it can also induce cancer cell death without affecting normal cells.
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- Vuppalapati, Karuna K, et al.
(författare)
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Targeted Deletion of Autophagy Genes Atg5 or Atg7 in the Chondrocytes Promotes Caspase-Dependent Cell Death and Leads to Mild Growth Retardation.
- 2015
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Ingår i: Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research. - : Wiley. - 1523-4681. ; 30:12, s. 2249-2261
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Tidskriftsartikel (refereegranskat)abstract
- Longitudinal bone growth takes place in epiphyseal growth plates located in the ends of long bones. The growth plate consists of chondrocytes traversing from the undifferentiated (resting zone) to the terminally differentiated (hypertrophic zone) stage. Autophagy is an intracellular catabolic process of lysosome-dependent recycling of intracellular organelles and protein complexes. Autophagy is activated during nutritionally depleted or hypoxic conditions in order to facilitate cell survival. Chondrocytes in the middle of the growth plate are hypoxic and nutritionally depleted owing to the avascular nature of the growth plate. Accordingly, autophagy may facilitate their survival. To explore the role of autophagy in chondrocyte survival and constitutional bone growth, we generated mice with cartilage-specific ablation of either Atg5 (Atg5cKO) or Atg7 (Atg7cKO) by crossing Atg5 or Atg7 floxed mice with cartilage-specific collagen type 2 promoter-driven Cre. Both Atg5cKO and Atg7cKO mice showed growth retardation associated with enhanced chondrocyte cell death and decreased cell proliferation. Similarly, inhibition of autophagy by Bafilomycin A1 (Baf) or 3-methyladenine (3MA) promoted cell death in cultured slices of human growth plate tissue. To delineate the underlying mechanisms we employed ex vivo cultures of mouse metatarsal bones and RCJ3.IC5.18 rat chondrogenic cell line. Baf or 3MA impaired metatarsal bone growth associated with processing of caspase-3 and massive cell death. Similarly, treatment of RCJ3.IC5.18 chondrogenic cells by Baf also showed massive cell death and caspase-3 cleavage. This was associated with activation of caspase-9 and cytochrome C release. Altogether, our data suggest that autophagy is important for chondrocyte survival, and inhibition of this process leads to stunted growth and caspase-dependent death of chondrocytes. © 2015 American Society for Bone and Mineral Research.
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