Abstract

Protein tyrosine phosphatase type IVA 3 (PTP4A3) is implicated in the metastasis of hepatocellular carcinoma (HCC), yet its precise regulatory mechanisms remain elusive. This study aimed to identify the interacting substrates and underlying mechanisms of PTP4A3 to inform the optimization of PTP4A3-targeted therapies in HCC. Using the TurboID system and protein mass spectrometry, over 500 candidate substrates for PTP4A3 were identified. A direct interaction between PTP4A3 and epidermal growth factor receptor (EGFR) was also validated. Ti-IMAC protein phosphorylation mass spectrometry revealed that PTP4A3 dephosphorylates Thr693 of EGFR. This dephosphorylation event enhanced EGFR-ERBB2 heterodimerization, leading to increased RhoA-GTPase activation, cell motility, and cytoskeletal rearrangement. EGFR/ERBB2 inhibition effectively suppressed PTP4A3-driven metastasis both in vitro and in vivo. This study reveals a novel mechanism by which PTP4A3 promotes HCC metastasis via direct dephosphorylation of EGFR at Thr693. Lapatinib, an inhibitor of EGFR/ERBB2, effectively suppressed PTP4A3-driven HCC metastasis, offering a potential therapeutic strategy for HCC patients with high PTP4A3 expression. The regulatory mechanism of PTP4A3 regulating HCC cell metastasis . PTP4A3 overexpression promotes EGFR-ERBB2 heterodimer formation by dephosphorylating EGFR at Thr693, inhibiting its degradation. This sustained EGFR activation enhances HCC cell motility via RhoA-GTP signaling. Lapatinib, an EGFR/ERBB2 inhibitor, effectively counteracts the pro-migratory effects of PTP4A3.