Targeting DDX5 Inhibitors to Battle Cancers & Viral Infections

DDX5 (Dead-Box 5 or p68) is a member of a family of 37 “DEAD-Box” ATP-dependent RNA helicases that play a role in nearly all aspects of RNA processing and also act as nucleic acid recognition receptors for viral immunity. Dysregulation and/or overexpression of DDX5 can lead to tumor progression. It can also be hijacked by some viruses, such as SARS-CoV-2, to enable their replication.^3,4

Structure and Function

DEAD-Box helicases are named for the core Asp-Glu-Ala-Asp (DEAD) amino acid sequence they share. Members of this family also possess highly conserved ATPase and helicase domains, flanked by individually distinctive N and/or C-terminal motifs. These distinctive flanking sequences inform the particular protein-protein and RNA-protein interactions for the diverse human DEAD-Box proteins.

DDX5 is a 614 amino acid protein consisting of an N-terminal Q motif, an ATP binding Rec A-like domain, a hinge region, a Rec A-like helicase domain, and a C-terminal RGS-RGG motif. The Q motif is necessary for single-stranded RNA binding and for proper conformational dynamics in response to ATP binding and hydrolysis. While both the ATP binding domain and the helicase domain share and cooperate in conformationally dictated ATP binding and hydrolysis, RNA binding, and helicase functions, the N-terminal Rec A-like domain principally orchestrates ATP binding and hydrolysis, while the C-terminal Rec A-like domain mainly controls RNA binding and helicase activities. The C-terminal RGS-RGG (Arg-Gly-Ser-Arg-Gly-Gly) motif modulates DDX5-specific biological functions through its interaction with RNA and DDX3. More distal C-terminal motifs have been proposed (such as IQ, SH2, and c-Abl) that are adding more complexity to DDX5’s repertoire of protein interactions.¹

While DDX5 is predominantly a nuclear protein, it can shuttle between the nucleus and the cytoplasm, depending upon cell cycle status, cell type, and given post-translational modification. Indeed, c-Abl mediated phosphorylation of DDX5’s tyrosine 593 promotes its nuclear export. DDX5’s activities are vast, varied, and expanding. It functions as an RNA chaperone, a transcriptional regulator, a facilitator of mRNA processing, an enabler of nonsense-mediated decay, an important player in R loop resolution, an aid in micro-RNA processing, and an instrument of ribosomal biogenesis.¹

Implications in Cancers and Viral Infections

DDX5 potentiates important signaling pathways, such as Wnt, Akt, and mTOR, and even upregulates glucose metabolism²; not surprisingly, dysregulation can affect cell proliferation, cell cycle control, and tumorigenesis. DDX5 overexpression correlates to many cancers, including breast, colorectal, gastric, lung, endometrial, and prostate. It has also been found to interact with micro RNAs and long noncoding RNAs (lcnRNAs) to potentiate their activities and drive cancer progression. In colon cancer, DDX5 and its paralog, DDX17, bind to beta-catenin to activate the transcription of various potent oncogenes, such as c-Myc and cyclin D1.³

The role of DDX5 in viral defense and disease is ambiguous. Like several other DDXs, most notably DDX3, it acts as a viral RNA sensor and contributes to immunity via type I interferon induction, but it is also hijacked by some viruses to enable their propagation. For example, DDX5 inhibits myxoma virus replication, represses hepatitis B virus (HBV) minichromosomal function, and enables STAT1 signaling/the IFN innate immune response in HBV-infected cells.⁴ On the other hand, DDX5 binds the SARS-CoV helicase to enhance viral proliferation, facilitates HIV-1 mRNA export, and enhances influenza virus polymerase activity. ⁴

Discovery of DDX5 Inhibitors to Delineate the Complex Roles

Development of DDX5 inhibitors will help to delineate the complex roles of DDX5 and other DEAD-Box helicase in cancer and viral infection and hopefully lead to development of improved therapeutic strategies. Use of the Transcreener^® ADP² assay for detection of helicase-linked ATP hydrolysis is a robust, HTS-proven method for identifying and characterizing DEAD-Box helicase inhibitors. Our DDX3 application provides an excellent example of using this assay to study DEAD-Box helicases, like DDX5.

View the DDX3 Application Assay

References

1. Xing, Z. et al. (2019) The DDX5/Dbp2 subfamily of DEAD-box RNA helicases. Wiley Interdiscip Rev RNA. 10(2): e1519. Review. https://doi.org/10.1002/wrna.1519.
2. Xing, Z. et al. (2017) Characterization of the mammalian DEAD-box protein DDX5 reveals functional conservation with S. cerevisiae ortholog Dbp2 in transcriptional control and glucose metabolism. RNA. 23: 1125-1138. Review. https://doi.org/10.1261/rna.060335.116.
3. Li, F. et al. (2021) Multiple functions of the DEAD-box RNA helicase, DDX5 (p68), make DDX5 a superior oncogenic biomarker and target for targeted cancer therapy. Am J Cancer Res. 11(10):5190-5213. Review. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8569338/?report=classic.
4. Cheng, W. et al. (2018) DDX5 RNA Helicases: Emerging Roles in Viral Infection. Int. J. Mol. Sci. 19(4), 1122. Review. https://doi.org/10.3390/ijms19041122.