A Novel Isoform of TUCAN Is Overexpressed in Human Cancer Tissues and Suppresses Both Caspase-8– and Caspase-9–Mediated Apoptosis

Abstract

Caspase-associated recruitment domains (CARD) are proteinprotein interaction modules found extensively in proteins that play important roles in apoptosis. One of the CARD-containing proteins, TUCAN (CARD8), was reported previously as an antiapoptotic protein with a molecular weight of 48 kDa, which was up-regulated in colon cancer cells. We identified a novel isoform of TUCAN with a molecular weight of 54 kDa. The new variant of TUCAN, termed TUCAN-54, was expressed in gastric, colon, and breast cancer tissues but was barely detected in normal noncancerous tissues, whereas 48-kDa TUCAN was detected in tumor tissues and noncancerous tissues. To know the function of TUCAN-54 in the apoptosis of cancer cells, TUCAN-54 was overexpressed in tumor cells by gene transfection. Its overexpression inhibited pro-caspase-9 activation, leading to the suppression of the cell death induced by a protein kinase inhibitor, staurosporine, or a chemotherapeutic reagent, etoposide (VP-16). In contrast, specific small interfering RNA-mediated suppression of TUCAN-54 expression in tumor cells increased the VP-16-induced cell death rate, indicating that expression of TUCAN-54 might be associated with chemoresistance of tumor cells. In addition, it inhibited caspase-8 activation as well, thereby suppressing Fas-induced cell death. It was revealed that Fas-associated death domain was physically associated with TUCAN-54 but not with 48-kDa TUCAN. Thus, TUCAN-54 might be a novel tumor-specific antiapoptotic molecule expressed in a variety of human cancer tissues, which might aggravate malignant potential of cancer cells, such as chemoresistance and immunoresistance.

Association of CARD8 with inflammatory bowel disease in Koreans

“…[13][14][15][16] CARD8 acts as a negative regulator of nuclear factor-kappa B 13,14 and is a suppressor of apoptosis. 15,16 Recent studies found that CARD8 interacted with NALP3 (NACHT; NAIP, CIIA, HET-E, and TP1; Neuronal apoptosis inhibitor protein (NAIP), MHC Class II transcription activator (CIIA), Incompatibility locus protein from Podospora anserina (HET-E), and Telomerase-associated protein (TP1) domain, leucine-rich repeat and pyrin domain-containing protein 3) and NOD2 (nucleotide-binding oligomerization domain 2), members of the NOD-like receptor family, implying a role for CARD8 in the regulation of multiple inflammatory pathways. 17,18 Tumor necrosis factor-a, interleukin-1b (IL-1b), and IL-6 are characteristically overproduced in CD patients.…”

A Prickly Subject: Apoptotic Regulation by Hedgehog Morphogens

“…Through a homophilic CARD-CARD interaction, the smaller form binds Caspase-9 and blocks a variety of intrinsic-pathway activating insults [88,89]. In contrast the 54kDa, which contains a unique amino-terminus absent in 48kDa form, binds FADD and blocks FasL-and extrinsic Caspase-8-mediated apoptosis [89].…”

Novel isoforms of the CARD8 (TUCAN) gene evade a nonsense mutation

“…The TUCAN/CARD8 antibody was raised against residues 99 -115 of T54 (encoded by part of exon 7). 8 The membrane was washed in PBS and incubated for 1 h at 41C in PBS containing 5% non-fat milk and anti-mouse IgG antibody (Dako, Stockholm, Sweden), washed in PBS and incubated with the ECL detection kit (Amersham Biosciences). Chemiluminescence was visualised using Hyperfilm ECL (Amersham Biosciences).…”

“…2 -4 Another CARD-containing protein, CARD8, also named TUCAN (tumour-upregulated CARD-containing antagonist of caspase-9) or CARDINAL, is implicated in the regulation of both apoptosis and inflammation, and is known to undergo differential splicing leading to at least two protein isoforms. 5 -8 A 48 kDa isoform (T48) expressed mainly in monocytes, placenta, lymph nodes and spleen 8 interacts directly with caspase-1 and can induce apoptosis, whereas a larger 54 kDa isoform (T54) is overexpressed in some cancers and suppresses caspase-mediated apoptosis. 8 CARD8 also interacts with IKKg/NEMO, leading to inhibition of NF-kB activity.…”

“…5 -8 A 48 kDa isoform (T48) expressed mainly in monocytes, placenta, lymph nodes and spleen 8 interacts directly with caspase-1 and can induce apoptosis, whereas a larger 54 kDa isoform (T54) is overexpressed in some cancers and suppresses caspase-mediated apoptosis. 8 CARD8 also interacts with IKKg/NEMO, leading to inhibition of NF-kB activity. 7 Its gene maps to a region that has shown linkage to inflammatory bowel disease (IBD) in several genome-wide linkage scans, 9 -12 confirmed in a metaanalysis of 10 genome-wide linkage studies.…”