DDX3

Oncogenic intercellular signaling is regulated by extracellular vesicles (EVs), but the underlying mechanisms remain mostly unclear. Since TCTP (translationally controlled tumor protein) is an EV component, we investigated whether it has a role in genotoxic stress signaling and malignant transformation. By generating a Tctp-inducible knockout mouse model (Tctp-/f- ), we report that Tctp is required for genotoxic stress-induced apoptosis signaling via small EVs (sEVs). Human breast cancer cells knocked-down for TCTP show impaired spontaneous EV secretion, thereby reducing sEV-dependent malignant growth...

Programmed death-1 (PD-1)/PD ligand-1 (PD-L1)-mediated immune escape contributes to cancer development and has been targeted as an anti-cancer strategy. Here, we show that inhibition of the RNA helicase DDX3 increased CD8+ T cell infiltration in syngeneic oral squamous cell carcinoma tumors. DDX3 knockdown compromised interferon-γ-induced PD-L1 expression and, in particular, reduced the level of cell-surface PD-L1. DDX3 promoted surface PD-L1 expression by recruiting the adaptor protein 2 (AP2) complex to the 3' UTR of PD-L1 mRNA...

Upregulation of nitric oxide (NO) production contributes to the pathogenesis of numerous diseases via S-nitrosylation, a post-translational modification of proteins. This process occurs due to the oxidative reaction between NO and a cysteine thiol group; however, the extent of this reaction remains unknown. S-Nitrosylation of PRMT1, a major asymmetric arginine methyltransferase of histones and numerous RNA metabolic proteins, was induced by NO donor treatment. We found that nitrosative stress leads to S-nitrosylation of cysteine 119, located near the active site, and attenuates the enzymatic activity of PRMT1...

We previously demonstrated that RNA helicase DDX3X (DDX3) can be a therapeutic target in Ewing sarcoma (EWS), but its role in EWS biology remains unclear. The present work demonstrates that DDX3 plays a unique role in DNA damage repair (DDR). We show that DDX3 interacts with several proteins involved in homologous recombination, including RAD51, RECQL1, RPA32, and XRCC2. In particular, DDX3 colocalizes with RAD51 and RNA:DNA hybrid structures in the cytoplasm of EWS cells. Inhibition of DDX3 RNA helicase activity increases cytoplasmic RNA:DNA hybrids, sequestering RAD51 in the cytoplasm, which impairs nuclear translocation of RAD51 to sites of double-stranded DNA breaks, thus increasing sensitivity of EWS to radiation treatment, both in vitro and in vivo ...

BACKGROUND: DDX3 is a protein with RNA helicase activity that is involved in a variety of biological processes, and it is an important protein target for the development of broad-spectrum antiviral drugs, multiple cancers and chronic inflammation. OBJECTIVE: The objective of this study is to establish a simple and efficient method to express and purify DDX3 protein in E. coli, and the recombinant DDX3 should maintain helicase activity for further tailor-made screening and biochemical function validation...

Necroptosis is a regulated necrotic cell death mechanism and plays a crucial role in the progression of cancers. However, the potential role and mechanism of necroptosis in colorectal cancer (CRC) has not been fully elucidated. In this study, we found that nuclear receptor subfamily 4 group A member 1 (NR4A1) was highly expressed in CRC cells treated with TNF-α, Smac mimetic, and z-VAD-FMK (TSZ). The depletion of NR4A1 significantly enhanced the sensitivity of CRC cells to TSZ-induced necroptosis, while NR4A1 overexpression suppressed these effects, as evidenced by the LDH assay, flow cytometry analysis of cell death, PI staining, and expression analysis of necrosome complexes (RIPK1, RIPK3, and MLKL)...

HIV-1 continues to be a major global health challenge. Current HIV-1 treatments are effective but need lifelong adherence. An HIV-1 cure should eliminate the latent viral reservoir that persists in people living with HIV-1. Different methods have been investigated that focus on reactivation and subsequent elimination of the HIV-1 reservoir, and it is becoming clear that a combination of compounds with different mechanisms of actions might be more effective. Here, we target two host factors, inhibitor of apoptosis proteins that control apoptosis and the DEAD-box helicase DDX3, facilitating HIV mRNA transport/translation...

Interferon-stimulated genes (ISGs) are induced in response to interferon expression due to viral infections. Role of these ISGs can be variable in different cells or organs. Our study highlights such cell-specific role of an ISG, Ddx3, which regulates the translation of mRNAs essential for interferon induction (PACT) and interferon signaling (STAT1) in a cell-specific manner. Our study also highlights the role of PACT in RNA virus-induced RLR signaling. Our study depicts how Ddx3 regulates innate immune signaling pathways in an indirect manner...

Membraneless organelles, or biomolecular condensates, enable cells to compartmentalize material and processes into unique biochemical environments. While specific, attractive molecular interactions are known to stabilize biomolecular condensates, repulsive interactions, and the balance between these opposing forces, are largely unexplored. Here, we demonstrate that repulsive and attractive electrostatic interactions regulate condensate stability, internal mobility, interfaces, and selective partitioning of molecules both in vitro and in cells...

DEAD-box RNA helicases are ATP-dependent RNA binding proteins and RNA-dependent ATPases that possess weak, nonprocessive unwinding activity in vitro, but they can form long-lived complexes on RNAs when the ATPase activity is inhibited. Ded1 is a yeast DEAD-box protein, the functional ortholog of mammalian DDX3, that is considered important for the scanning efficiency of the 48S pre-initiation complex ribosomes to the AUG start codon. We used a modified PAR-CLIP technique, which we call quicktime PAR-CLIP (qtPAR-CLIP), to crosslink Ded1 to 4-thiouridine-incorporated RNAs in vivo using UV light centered at 365 nm...

Human papillomavirus (HPV) infection is a causative agent of cervical cancer (CC). N6-methyladenosine (m6A) modification is implicated in carcinogenesis and tumor progression. However, the involvement of m6A modification in HPV-involved CC remains unclear. Here we showed that HPV E6/7 oncoproteins affected the global m6A modification and E7 specifically promoted the expression of ALKBH5. We found that ALKBH5 was significantly upregulated in CC and might serve as a valuable prognostic marker. Forced expression of ALKBH5 enhanced the malignant phenotypes of CC cells...

Long noncoding RNAs (lncRNAs) play pivotal roles in tumor development. To identify dysregulated lncRNAs in gastric cancer (GC), we analyzed genome-wide trimethylation of histone H3 lysine 4 (H3K4me3) to screen for transcriptionally active lncRNA genes in the non-tumorous gastric mucosa of patients with GC and healthy individuals. We found that H3K4me3 at TM4SF1-AS1 was specifically upregulated in GC patients and that the expression of TM4SF1-AS1 was significantly elevated in primary and cultured GC cells. TM4SF1-AS1 contributes to GC cell growth in vitro and in vivo, and its oncogenic function is mediated, at least in part, through interactions with purine-rich element-binding protein α (Pur-α) and Y-box binding protein 1 (YB-1)...

We previously demonstrated that RNA helicase DDX3X (DDX3) can be a therapeutic target in Ewing sarcoma (EWS), but its role in EWS biology remains unclear. The present work demonstrates that DDX3 plays a unique role in DNA damage repair (DDR). We show that DDX3 interacts with several proteins involved in homologous recombination, including RAD51, RECQL1, RPA32, and XRCC2. In particular, DDX3 colocalizes with RAD51 and RNA:DNA hybrid structures in the cytoplasm of EWS cells. Inhibition of DDX3 RNA helicase activity increases cytoplasmic RNA:DNA hybrids, sequestering RAD51 in the cytoplasm, which impairs nuclear translocation of RAD51 to sites of double-stranded DNA breaks thus increasing sensitivity of EWS to radiation treatment, both in vitro and in vivo ...

The canonical DEAD-box helicase, eIF4A, unwinds 5' UTR secondary structures to promote mRNA translation initiation. Growing evidence has indicated that other helicases, such as DHX29 and DDX3/ded1p, also function to promote the scanning of the 40S subunit on highly structured mRNAs. It is unknown how the relative contributions of eIF4A and other helicases regulate duplex unwinding on an mRNA to promote initiation. Here, we have adapted a real-time fluorescent duplex unwinding assay to monitor precisely helicase activity in the 5' UTR of a reporter mRNA that can be translated in a cell-free extract in parallel...

Due to the lack of predictive biomarkers and the lack of conspicuous symptoms at the early stage, hepatocellular carcinoma (HCC) remains difficult to diagnose and treat effectively. During cancer development, exosomes secreted from tumor cells carry functional molecules to surrounding recipient cells, thereby participating in the regulation of cancer progression. DDX3, a DEAD-box RNA helicase, performs many important functions in several cellular processes and is therefore implicated as a tumor suppressor in HCC...

While protein synthesis is vital for the majority of cell types of the human body, diversely differentiated cells require specific translation regulation. This suggests the specialization of translation machinery across tissues and organs. Using transcriptomic data from GTEx, FANTOM, and Gene Atlas, we systematically explored the abundance of transcripts encoding translation factors and aminoacyl-tRNA synthetases (ARSases) in human tissues. We revised a few known and identified several novel translation-related genes exhibiting strict tissue-specific expression...

DDX3X is an RNA helicase with many functions in RNA metabolism such as mRNA translation, alternative pre-mRNA splicing and mRNA stability, but also plays a role as a regulator of transcription as well as in the Wnt/beta-catenin- and Nf-κB signaling pathways. The gene encoding DDX3X is located on the X-chromosome, but escapes X-inactivation. Hence females have two active copies and males only one. However, the Y chromosome contains the gene for the male DDX3 homologue, called DDX3Y , which has a very high sequence similarity and functional redundancy with DDX3X, but shows a more restricted protein expression pattern than DDX3X...

Renal cell carcinoma (RCC) is the most difficult-to-treat form of kidney cancer with a median 5-year survival of 10% under metastatic setting. In RCC, although cytoreductive nephrectomy is common, approximately 20-30% of patients will develop recurrent cancer after surgery, which highlights the need for an effective therapy. Rho-GTPases viz, Rac-1 and Cdc42 are the central regulators of cancer cell migration and invasion and thus metastasis in multiple cancer types. Hence, we elucidated the role of Ketorolac, a modulator Rho-GTPases against RCC through potentiation of tumor suppressor Par-4...

This study aimed to identify the direct pharmacological targets of Jingfang Granules in treating infectious pneumonia via "target fishing" strategy. Moreover, the molecular mechanism of Jingfang Granules in treating infectious pneumonia was also investigated based on target-related pharmacological signaling pathways. First, the Jingfang Granules extract-bound magnetic nanoparticles were prepared, which were incubated with lipopolysaccharide(LPS)-induced mouse pneumonia tissue lysates. The captured proteins were analyzed by high-resolution mass spectrometry(HRMS), and the target groups with specific binding to the Jingfang Granules extract were screened out...

OBJECTIVE: Cholangiocarcinoma (CCA) is a cancer of the bile duct with a poor prognosis. The present study examined the ability of curcumin to sensitize apoptosis in the TNF-related apoptosis-inducing ligand (TRAIL)-resistant CCA cell lines of HuCCA-1 and KKU-213A. METHODS: Apoptosis was measured using a TUNEL assay. Protein expression was determined by immunoblotting. Membrane death receptor 5 (DR5) was detected by flow cytometry. Protein complex was examined by co-immunoprecipitation...