Dataset Information

Identification of a novel anti-apoptotic E3 ubiquitin ligase that ubiquitinates antagonists of inhibitor of apoptosis proteins SMAC, HtrA2, and ARTS.

ABSTRACT: Identification of new anti-apoptotic genes is important for understanding the molecular mechanisms underlying apoptosis and tumorigenesis. The present study identified a novel anti-apoptotic gene named AREL1, which encodes a HECT (homologous to E6-AP carboxyl terminus) family E3 ubiquitin ligase. AREL1 interacted with and ubiquitinated IAP antagonists such as SMAC, HtrA2, and ARTS. However, AREL1 was cytosolic and did not localize to nuclei or mitochondria. The interactions between AREL1 and the IAP antagonists were specific for apoptosis-stimulated cells, in which the IAP antagonists were released into the cytosol from mitochondria. Furthermore, the ubiquitination and degradation of SMAC, HtrA2, and ARTS were significantly enhanced in AREL1-expressing cells following apoptotic stimulation, indicating that AREL1 binds to and ubiquitinates cytosolic but not mitochondria-associated forms of IAP antagonists. Furthermore, the anti-apoptotic role of AREL1-mediated degradation of SMAC, HtrA2, and ARTS was shown by simultaneous knockdown of three IAP antagonists, which caused the inhibition of caspase-3 cleavage, XIAP degradation, and induction of apoptosis. Therefore, the present study suggests that AREL1-mediated ubiquitination and degradation of cytosolic forms of three IAP antagonists plays an important role in the regulation of apoptosis.

SUBMITTER: Kim JB

PROVIDER: S-EPMC3636887 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature

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Identification of a novel anti-apoptotic E3 ubiquitin ligase that ubiquitinates antagonists of inhibitor of apoptosis proteins SMAC, HtrA2, and ARTS.

Kim Jung-Bin JB Kim So Youn SY Kim Byeong Mo BM Lee Hunjin H Kim Insook I Yun Jeanho J Jo Yejin Y Oh Taeheun T Jo Yongsam Y Chae Hee-Don HD Shin Deug Y DY

The Journal of biological chemistry 20130311 17

Identification of new anti-apoptotic genes is important for understanding the molecular mechanisms underlying apoptosis and tumorigenesis. The present study identified a novel anti-apoptotic gene named AREL1, which encodes a HECT (homologous to E6-AP carboxyl terminus) family E3 ubiquitin ligase. AREL1 interacted with and ubiquitinated IAP antagonists such as SMAC, HtrA2, and ARTS. However, AREL1 was cytosolic and did not localize to nuclei or mitochondria. The interactions between AREL1 and the ...[more]

PMID: 23479728

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Project description:BACKGROUND:Colorectal cancer (CRC) is the fourth leading cause of cancer-related deaths worldwide, and deciphering underlying molecular mechanism is essential. The loss of monoubiquitinated histone H2B (H2Bub1) was correlated with poor prognosis of CRC patients and, accordingly, H2Bub1 was suggested as a tumor-suppressive mark. Surprisingly, our previous work revealed that the H2B ubiquitin ligase RING finger protein 40 (RNF40) might exert tumor-promoting functions. Here, we investigated the effect of RNF40 loss on tumorigenic features of CRC cells and their survival in vitro. METHODS:We evaluated the effects of RNF40 depletion in several human CRC cell lines in vitro. To evaluate cell cycle progression, cells were stained with propidium iodide and analyzed by flow cytometry. In addition, to assess apoptosis rates, caspase 3/7 activity was assessed in a Celigo® S-based measurement and, additionally, an Annexin V assay was performed. Genomic occupancy of H2Bub1, H3K79me3, and H3K27ac was determined by chromatin immunoprecipitation. Transcriptome-wide effects of RNF40 loss were evaluated based on mRNA-seq results, qRT-PCR, and Western blot. To rescue apoptosis-related effects, cells were treated with Z-VAD-FMK. RESULTS:Human CRC cell lines displayed decreased cell numbers in vitro after RNF40 depletion. While the differences in confluence were not mediated by changes in cell cycle progression, we discovered highly increased apoptosis rates after RNF40 knockdown due to elevated caspase 3/7 activity. This effect can be explained by reduced mRNA levels of anti-apoptotic and upregulation of pro-apoptotic BCL2 family members. Moreover, the direct occupancy of the RNF40-mediated H2B monoubiquitination was observed in the transcribed region of anti-apoptotic genes. Caspase inhibition by Z-VAD-FMK treatment rescued apoptosis in RNF40-depleted cells. However, knockdown cells still displayed decreased tumorigenic features despite the absence of apoptosis. CONCLUSIONS:Our findings reveal that RNF40 is essential for maintaining tumorigenic features of CRC cells in vitro by controlling the expression of genes encoding central apoptotic regulators.

Dataset Information

Identification of a novel anti-apoptotic E3 ubiquitin ligase that ubiquitinates antagonists of inhibitor of apoptosis proteins SMAC, HtrA2, and ARTS.

Publications

Identification of a novel anti-apoptotic E3 ubiquitin ligase that ubiquitinates antagonists of inhibitor of apoptosis proteins SMAC, HtrA2, and ARTS.

Similar Datasets

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