Identification of TRIM25 as a Negative Regulator of Caspase-2 Expression Reveals a Novel Target for Sensitizing Colon Carcinoma Cells to Intrinsic Apoptosis.

Usman Nasrullah,Kristina Haeussler,Wolfgang Eberhardt,Josef Pfeilschifter,Abhiruchi Biyanee,Ilka Wittig

doi:10.3390/cells8121622

Usman Nasrullah, Kristina Haeussler + Show 4 more

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https://doi.org/10.3390/cells8121622

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Journal: Cells	Publication Date: Dec 12, 2019
Citations: 13	License type: CC BY 4.0

Affiliation: Goethe University Frankfurt, University of Münster

Abstract

Colorectal cancer (CRC) is one of the most common cancers that is characterized by a high mortality due to the strong metastatic potential of the primary tumor and the high rate of therapy resistance. Hereby, evasion of apoptosis is the primary underlying cause of reduced sensitivity of tumor cells to chemo- and radiotherapy. Using RNA affinity chromatography, we identified the tripartite motif-containing protein 25 (TRIM25) as a bona fide caspase-2 mRNA-binding protein in colon carcinoma cells. Loss-of-function and gain-of-function approaches revealed that TRIM25 attenuates the protein levels of caspase-2 without significantly affecting caspase-2 mRNA levels. In addition, experiments with cycloheximide revealed that TRIM25 does not affect the protein stability of caspase-2. Furthermore, silencing of TRIM25 induced a significant redistribution of caspase-2 transcripts from RNP particles to translational active polysomes, indicating that TRIM25 negatively interferes with caspase-2 translation. Functionally, the elevation in caspase-2 upon TRIM25 depletion significantly increased the sensitivity of colorectal cells to drug-induced intrinsic apoptosis as implicated by increased caspase-3 cleavage and cytochrome c release. Importantly, the apoptosis-sensitizing effects by transient TRIM25 knockdown were rescued by concomitant silencing of caspase-2, demonstrating a critical role of caspase-2. Inhibition of caspase-2 by TRIM25 implies a survival mechanism that critically contributes to chemotherapeutic drug resistance in CRC.

Highlights

The occurrence of drug resistance is a major obstacle that frequently limits the therapeutic benefit of cancer therapeutics
Since the negative regulation of caspase-2 by human antigen R (HuR) depends on the 50 untranslated region (50 UTR), for affinity purification, biotin-labelled in vitro-transcribed mRNAs encompassing either the 50 -UTR of caspase-2, or alternatively, the coding region of caspase-2 were used as baits
Among various eukaryotic translation initiation factors and some well-known RNA-binding proteins, including HuR, we identified the tripartite motif-containing protein (TRIM) 25, synonymously denoted as estrogen-responsive finger protein (Efp), as a protein strongly associated with the 50 UTR but only with a weak affinity to the cdr of caspase-2 mRNA (Supplementary Table S1)

Summary

Introduction

The occurrence of drug resistance is a major obstacle that frequently limits the therapeutic benefit of cancer therapeutics. For this reason, the implementation of novel therapies that resensitize tumor cells towards current tumor therapies is urgently needed. Therapy-resistant tumor cells are characterized by impaired activation of caspases, a family of cysteine-aspartate proteases that mediate the proteolytic processing of diverse downstream substrates in response to disruptive insults (for a review, see [2,5]). In addition to elevations in IAP proteins, we previously identified direct inhibition of caspase-2 translation by the ubiquitous mRNA-binding protein human antigen R (HuR) as a novel path of therapy resistance in colon carcinoma cells [6,7,8] (for a review, see [9]). Regardless, by acting as an initiator caspase, caspase-2 plays a critical role in the execution phase of apoptosis in response to DNA damage, genotoxic drugs, or mitotic catastrophe (for a review, see [10,11,12])

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