Context-specific roles of DDX60 in colorectal cancer via autophagy regulation and DDX58 signaling.

DEAD-box RNA helicases have emerged as critical regulators in cancer biology, yet the functional role of DDX60 in colorectal cancer (CRC) remains poorly characterized and tissue-context dependent. Existing reports suggest opposing roles of DDX60 across tumor types, potentially mediated by autophagy and innate immune signaling. Understanding the dual roles of DDX60 in CRC is crucial for delineating its therapeutic relevance. DExD/H-box helicase 60 (DDX60) is a newly identified member of the DEAD-box (DDX) RNA helicase family. Existing evidence suggests that the expression patterns and functions of DDX60 vary across tumor types in a tissue-specific manner. However, its precise biological role and underlying molecular mechanisms in colorectal cancer (CRC) remain unclear. The expression of DDX60 in CRC cell lines was examined by transcriptome sequencing (RNA-seq) and Western blot (WB) analysis. Lentiviral transduction was employed to achieve DDX60 overexpression (OE-DDX60) and knockdown (KD-DDX60) in both cellular assays and xenograft models. Cell proliferation, migration, and invasion were assessed using the Cell Counting Kit-8 (CCK-8), EdU incorporation, wound healing, and Transwell assays. Immunofluorescence staining was performed to detect microtubule-associated protein 1A/1B-light chain 3B (LC3B) and sequestosome 1 (p62/SQSTM1) levels. Co-immunoprecipitation (Co-IP) assays were conducted to evaluate the interaction between DDX60 and DExD/H-box helicase 58 (DDX58) Lipid peroxidation products and inflammatory cytokines were quantified using commercial assay kits, while mitochondrial membrane potential was assessed via JC-1 staining. SW620 cells were subcutaneously injected into the groin region of nude mice to establish xenograft tumors, and tumor weight and volume were monitored. Histological changes and apoptosis in tumor tissues were analyzed using hematoxylin and eosin (H&E) staining and flow cytometry. DDX60 is highly expressed in CRC cell lines. In vitro experiments revealed that OE-DDX60 significantly promoted the proliferation, migration, and invasion of SW620 cells while inhibiting apoptosis, confirming its pro-tumorigenic role. These effects were accompanied by reduced levels of lipid peroxidation products and pro-inflammatory cytokines. In contrast, in vivo experiments showed that OE-DDX60 suppressed tumor growth and enhanced apoptosis in tumor tissues, indicating an anti-tumor effect. This was associated with increased levels of inflammatory cytokines and enhanced DDX58 signaling, which contradicted the in vitro findings. Additionally, DDX60 markedly activated autophagy in both in vitro and in vivo settings. Notably, administration of the autophagy inhibitor CA-5f exerted anti-tumor effects on CRC. In summary, this study elucidates the biological functions of DDX60 in CRC and provides novel experimental evidence supporting its potential as a therapeutic biomarker.