Journal: Cell Death & Disease

Article Title: The LEF1/CYLD axis and cIAPs regulate RIP1 deubiquitination and trigger apoptosis in selenite-treated colorectal cancer cells

doi: 10.1038/cddis.2014.13

Figure Lengend Snippet: Selenite-induced degradation of cIAPs facilitated apoptosis by promoting DISC formation. ( a and b ) After cIAP silencing using an siRNA mixture containing sequences targeted to cIAP1 and cIAP2 for 24 h, HCT116 and SW480 CRC cells were treated with 10 μ M selenite and subjected to FACS and western blot analysis. ( c and d ) HCT116 and SW480 cells were transfected with a plasmid expressing cIAP1 before selenite treatment and were then subjected to FACS and western blot analysis. ( e ) Coimmunoprecipitation using a RIP1 antibody was performed after transfection of CRC cells with the cIAP1 plasmid, followed by incubation with K63 polyubiquitin chain-specific antibody, caspase-8 antibody and FADD antibody

Article Snippet: CYLD siRNA (5′-AAGUACCGAAGGGAAGUAUAGtt-3′), cIAP1 siRNA (5′-UCCUGACAACUGGAGAGAAtt-3′), cIAP2 siRNA (5′-GAUUCGUUCAGAGUCUAAAtt-3′), LEF1 siRNA (5′-GGAAAAGAUCUUCGCCGAG-3′), caspase-8 siRNA (5′-AACCUCGGGAUACUGUCUGA-3′) and non-targeting control siRNA were acquired from GenePharma (Shanghai, China).

Techniques: Western Blot, Transfection, Plasmid Preparation, Expressing, Incubation

Journal: Cell Death & Disease

Article Title: The LEF1/CYLD axis and cIAPs regulate RIP1 deubiquitination and trigger apoptosis in selenite-treated colorectal cancer cells

doi: 10.1038/cddis.2014.13

Figure Lengend Snippet: Selenite demonstrated antitumour activity in a colon cancer xenograft model by triggering apoptosis via the LEF1/CYLD/cIAPs/caspase-8 signalling pathway. ( a ) Fourteen days after inoculation with HCT116 cells, nude mice (seven per group) were injected with PBS or selenite (2 mg/kg/d). Tumour volumes were calculated at the indicated intervals. The data are presented as the mean±S.D. ( b ) Representative images and quantitative analysis of in situ labelling of apoptosis cells using TUNEL assay; original magnification, 10 × . Scale bar, 100 μ m. * P <0.05. ( c ) Western blot analysis of proteins extracted from tumour tissues using antibodies against related molecules as indicated. ( d ) Immunohistochemistry results of the colon xenograft animal model using antibodies against cleaved caspase-8, cleaved caspase-3, CYLD and cIAPs (detecting both cIAP1 and cIAP2). Scale bar, 50 μ m

Article Snippet: CYLD siRNA (5′-AAGUACCGAAGGGAAGUAUAGtt-3′), cIAP1 siRNA (5′-UCCUGACAACUGGAGAGAAtt-3′), cIAP2 siRNA (5′-GAUUCGUUCAGAGUCUAAAtt-3′), LEF1 siRNA (5′-GGAAAAGAUCUUCGCCGAG-3′), caspase-8 siRNA (5′-AACCUCGGGAUACUGUCUGA-3′) and non-targeting control siRNA were acquired from GenePharma (Shanghai, China).

Techniques: Activity Assay, Injection, In Situ, TUNEL Assay, Western Blot, Immunohistochemistry, Animal Model