The RNA-binding protein YTHDF3 affects gastric cancer cell migration and response to paclitaxel by regulating EZRIN.

Background: Gastric cancer (GC) is the fourth most common cause of cancer-related mortality and the fifth most common cancer worldwide. Despite efforts, the identification of biomarkers and new therapeutic approaches for GC remains elusive. Recent studies have begun to reveal the role of N6-adenosine methylation (m⁶A) in the regulation of gene expression.

Methods: The expression of the reader YT521-B homology domain-containing family 3 (YTHDF3) in GC was assessed in 331 patients using immunohistochemistry. GC cell lines depleted of YTHDF3 using CRISPR-Cas9 were evaluated for migration, metastasis, orientation of the mitotic spindle, and response to paclitaxel. The association between YTHDF3 and EZRIN (EZR) mRNA was shown using RNA sequencing, immunofluorescence, real-time PCR, and RNA immunoprecipitation. The single-base elongation- and ligation-based qPCR amplification (SELECT) method was used to map m⁶A in the EZR transcript.

Results: YTHDF3 was significantly overexpressed in GC, and high levels of YTHDF3 were predictive of the response to chemotherapy. In GC cell lines, YTHDF3 was the most highly expressed reader protein. YTHDF3 depletion impaired cytoskeleton organization, cell migration and metastasis, and orientation of the mitotic spindle, leading to an increased response to paclitaxel. EZR was one of the downregulated targets in the YTHDF3 knockout cell models and was associated with the observed phenotype.

Conclusion: YTHDF3 contributes to cell motility and response to paclitaxel in GC cell lines, at least in part through EZR regulation. The YTHDF3-EZR regulatory axis is a novel molecular player in GC, with clinical relevance and potential therapeutic utility.