Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a cytokine that can trigger apoptosis in many types of human cancer cells via engagement of its two pro-apoptotic receptors TRAIL-R1 (DR4) and TRAIL-R2 (DR5). TRAIL can also activate several other signaling pathways such as activation of stress kinases, canonical NF-κB signaling and necroptosis. Though both receptors are ubiquitously expressed, their relative participation in TRAIL-induced signaling is still largely unknown. To analyze TRAIL receptor-specific signaling, we prepared Strep-tagged, trimerized variants of recombinant human TRAIL with high affinity for either DR4 or DR5 receptor. Using these receptor-specific ligands, we examined the contribution of individual pro-apoptotic receptors to TRAIL-induced signaling pathways. We found that in TRAIL-resistant colorectal HT-29 cells but not in pancreatic PANC-1 cancer cells, DISC formation and initial caspase-8 processing proceeds comparably via both DR4- and DR5-activated signaling. TRAIL-induced apoptosis, enhanced by the inhibitor of the Bcl-2 family ABT-737, or by the translation inhibitor homoharringtonine, proceeded in both cell lines predominantly via the DR5 receptor. ShRNA-mediated downregulation of DR4 or DR5 receptors in HT-29 cells also pointed to a stronger contribution of DR5 in TRAIL-induced apoptosis. In contrast to apoptosis, necroptotic signaling was activated similarly by both DR4- or DR5-specific ligands. Activation of auxiliary signaling pathways involving NF-κB or stress kinases proceeded under apoptotic conditions mainly in a DR5-dependent manner, while these signaling pathways were during necroptosis similarly activated by either of these ligands. Our study provides the first systematic insight into DR4-/DR5-specific signaling in colorectal and pancreatic cancer cells.

• Klíčová slova
• Apoptosis, Cancer, Necroptosis, Receptor-specific signaling, TRAIL,
• MeSH
• apoptóza genetika   MeSH
• buňky HT-29 MeSH
• kaspasa 8 genetika   MeSH
• kolorektální nádory genetika   patologie   MeSH
• lidé MeSH
• malá interferující RNA MeSH
• nádory slinivky břišní genetika   patologie   MeSH
• nekróza genetika   patologie   MeSH
• NF-kappa B genetika   MeSH
• pankreas metabolismus   patologie   MeSH
• proliferace buněk genetika   MeSH
• protein TRAIL genetika   MeSH
• regulace genové exprese u nádorů MeSH
• signální transdukce genetika   MeSH
• TRAIL receptory genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kaspasa 8 MeSH
• malá interferující RNA MeSH
• NF-kappa B MeSH
• protein TRAIL MeSH
• TNFRSF10A protein, human MeSH Prohlížeč
• TNFRSF10B protein, human MeSH Prohlížeč
• TNFSF10 protein, human MeSH Prohlížeč
• TRAIL receptory MeSH

Multifunctional adapter and chaperone protein Daxx participates in the regulation of a number of mainly transcription-related processes. Most notably in a complex with chromatin-remodelling ATPase ATRX, Daxx serves as a histone H3.3 chaperone at telomeric regions and certain genes. In this report we document that Daxx interacts with another chromatin-remodelling, ATPase Brg1. We confirm the Daxx-Brg1 association both in vitro and in cells and show that Daxx interacts with Brg1 in high-molecular-weight complexes. Ectopic co-expression of Daxx with Brg1 and PML could shift disperse nuclear localisation of Brg1 into PML bodies. Mapping the Daxx-Brg1 interaction revealed that Daxx preferentially binds the region between Brg1 N-terminal QLQ and HSA domains, but also weakly interacts with its C-terminal part. Brg1 interacted with both the central and N-terminal parts of Daxx. SiRNA-mediated down-regulation of Daxx in SW13 adrenal carcinoma cells markedly enhanced expression of Brg1-activated genes CD44 or SCEL, suggesting that Daxx either directly through Brg1 and/or indirectly via other factors is a negative regulator of their transcription. Our findings point to Brg1 as another chromatin-remodelling protein that might similarly, as ATRX, target Daxx to specific chromatin regions where it can carry out its chromatin- and transcription-regulating functions.

• Klíčová slova
• Adapter, Chromatin remodelling, Interaction, Transcriptional repression,
• Publikační typ
• časopisecké články MeSH

PURPOSE: To investigate the roles of BCL2, MCL1, and BCL-XL in the survival of diffuse large B-cell lymphoma (DLBCL). EXPERIMENTAL DESIGNS: Immunohistochemical analysis of 105 primary DLBCL samples, and Western blot analysis of 18 DLBCL cell lines for the expression of BCL2, MCL1, and BCL-XL. Pharmacologic targeting of BCL2, MCL1, and BCL-XL with ABT-199, homoharringtonine (HHT), and ABT-737. Analysis of DLBCL clones with manipulated expressions of BCL2, MCL1, and BCL-XL. Immunoprecipitation of MCL1 complexes in selected DLBCL cell lines. Experimental therapy aimed at inhibition of BCL2 and MCL1 using ABT-199 and HHT, single agent, or in combination, in vitro and in vivo on primary cell-based murine xenograft models of DLBCL. RESULTS: By the pharmacologic targeting of BCL2, MCL1, and BCL-XL, we demonstrated that DLBCL can be divided into BCL2-dependent and MCL1-dependent subgroups with a less pronounced role left for BCL-XL. Derived DLBCL clones with manipulated expressions of BCL2, MCL1, and BCL-XL, as well as the immunoprecipitation experiments, which analyzed MCL1 protein complexes, confirmed these findings at the molecular level. We demonstrated that concurrent inhibition of BCL2 and MCL1 with ABT-199 and HHT induced significant synthetic lethality in most BCL2-expressing DLBCL cell lines. The marked cytotoxic synergy between ABT-199 and HHT was also confirmed in vivo using primary cell-based murine xenograft models of DLBCL. CONCLUSIONS: As homoharringtonine is a clinically approved antileukemia drug, and ABT-199 is in advanced phases of diverse clinical trials, our data might have direct implications for novel concepts of early clinical trials in patients with aggressive DLBCL.

• MeSH
• apoptóza účinky léků   MeSH
• bicyklické sloučeniny heterocyklické aplikace a dávkování   MeSH
• bifenylové sloučeniny aplikace a dávkování   MeSH
• difúzní velkobuněčný B-lymfom klasifikace   farmakoterapie   genetika   patologie   MeSH
• harringtoniny aplikace a dávkování   MeSH
• homoharringtonin MeSH
• lidé MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nitrofenoly aplikace a dávkování   MeSH
• piperaziny aplikace a dávkování   MeSH
• proliferace buněk účinky léků   MeSH
• protein bcl-X biosyntéza   MeSH
• protein MCL-1 biosyntéza   genetika   MeSH
• protoonkogenní proteiny c-bcl-2 biosyntéza   genetika   MeSH
• regulace genové exprese u nádorů účinky léků   MeSH
• sulfonamidy aplikace a dávkování   MeSH
• xenogenní modely - testy antitumorózní aktivity MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ABT-737 MeSH Prohlížeč
• BCL2L1 protein, human MeSH Prohlížeč
• bicyklické sloučeniny heterocyklické MeSH
• bifenylové sloučeniny MeSH
• harringtoniny MeSH
• homoharringtonin MeSH
• MCL1 protein, human MeSH Prohlížeč
• nitrofenoly MeSH
• piperaziny MeSH
• protein bcl-X MeSH
• protein MCL-1 MeSH
• protoonkogenní proteiny c-bcl-2 MeSH
• sulfonamidy MeSH
• venetoclax MeSH Prohlížeč

Tumour necrosis factor (TNF) related apoptosis inducing ligand (TRAIL), a membrane-bound ligand from the TNF family, has attracted significant attention due to its rather specific and effective ability to induce apoptotic death in various types of cancer cells via binding to and activating its pro-apoptotic death receptors. However, a significant number of primary cancer cells often develop resistance to TRAIL treatment, and the signalling platform behind this phenomenon is not fully understood. Upon blocking endosomal acidification by the vacuolar ATPase (V-ATPase) inhibitors bafilomycin A1 (BafA1) or concanamycin A, we observed a significantly reduced initial sensitivity of several, mainly colorectal, tumour cell lines to TRAIL-induced apoptosis. In cells pretreated with these inhibitors, the TRAIL-induced processing of caspase-8 and the aggregation and trafficking of the TRAIL receptor complexes were temporarily attenuated. Nuclear factor κB or mitogen activated protein/stress kinase signalling from the activated TRAIL receptors remained unchanged, and neither possible lysosomal permeabilization nor acid sphingomyelinase was involved in this process. The cell surface expression of TRAIL receptors and their TRAIL-induced internalization were not affected by V-ATPase inhibitors. The inhibitory effect of BafA1, however, was blunted by knockdown of the caspase-8 inhibitor cFLIP. Altogether, the data obtained provide the first evidence that endosomal acidification could represent an important regulatory node in the proximal part of TRAIL-induced pro-apoptotic signalling.

• Klíčová slova
• TRAIL, V-ATPase, acidification, apoptosis, caspase-8,
• MeSH
• aktivace enzymů MeSH
• antitumorózní látky farmakologie   MeSH
• apoptóza MeSH
• down regulace MeSH
• endozomy metabolismus   MeSH
• FLIP (buněčný) metabolismus   MeSH
• kaspasa 8 metabolismus   MeSH
• koncentrace vodíkových iontů MeSH
• lidé MeSH
• makrolidy farmakologie   MeSH
• nádorové buněčné linie MeSH
• protein TRAIL farmakologie   MeSH
• sfingolipidy fyziologie   MeSH
• sfingomyelinfosfodiesterasa metabolismus   MeSH
• signální adaptorové proteiny receptorové domény smrti metabolismus   MeSH
• signální transdukce účinky léků   MeSH
• TRAIL receptory metabolismus   MeSH
• transport proteinů MeSH
• vakuolární protonové ATPasy antagonisté a inhibitory   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antitumorózní látky MeSH
• bafilomycin A1 MeSH Prohlížeč
• CASP8 protein, human MeSH Prohlížeč
• concanamycin A MeSH Prohlížeč
• FLIP (buněčný) MeSH
• kaspasa 8 MeSH
• makrolidy MeSH
• protein TRAIL MeSH
• sfingolipidy MeSH
• sfingomyelinfosfodiesterasa MeSH
• signální adaptorové proteiny receptorové domény smrti MeSH
• TNFSF10 protein, human MeSH Prohlížeč
• TRAIL receptory MeSH
• vakuolární protonové ATPasy MeSH