CircPAK1 Promotes the Progression of Hepatocellular Carcinoma Via Modulation of YAP Nucleus Localization by Interacting with 14-3-3ζ

Background: Circular RNA (circRNA), a new class of non-coding RNA, has obvious correlations with the occurrence and development of many diseases, including tumors. This study aimed to investigate the potential roles of circPAK1 in hepatocellular carcinoma (HCC).

Methods: High-throughput sequencing was performed on 3 pairs of HCC and matched normal tissues to determine the upregulated circRNAs. The expression level of circPAK1 was detected by qRT-PCR in HCC and paired with normal liver tissue samples. The effects of circPAK1 on proliferation, invasion, metastasis and apoptosis of HCC cells were evaluated by in vitro and in vivo experiments. We also constructed Chitosan/si-circPAK1 (CS/si-circPAK1) nanocomplexes using Chitosan material to evaluate its in vivo therapeutic effect on HCC. High-throughput sequencing, RNA-sequencing, RNA probe pull-down, RNA immunoprecipitation and Co-Immunoprecipitation assays were performed to explore the relationship between circPAK1, 14-3-3ζ, p-LATS1 and YAP. Exosomes isolated from lenvatinib-resistant HCC cell lines were used to evaluate the relationship between exosomal circPAK1 and lenvatinib resistance.

Results: CircPAK1, a novel circRNA, is highly expressed in HCC tumor tissues and cell lines as well as correlated with poor outcomes in HCC patients. Functionally, circPAK1 knockdown inhibited HCC cell proliferation, migration, invasion and angiogenesis while circPAK1 overexpression promoted HCC progression. The tumor-promoting phenotypes of circPAK1 on HCC were also confirmed by animal experiments. Importantly, the application of CS/si-circPAK1 nanocomplexes showed a better therapeutic effect on tumor growth and metastasis. Mechanistically, circPAK1 enhanced HCC progression by inactivating the Hippo signaling pathway, and this kind of inactivation is based on its competitively binding of 14-3-3 ζ with YAP, which weakens the recruitment and cytoplasmic fixation of 14-3-3 ζ to YAP, thus promoting YAP nucleus localization. Additionally, circPAK1 could be transported by exosomes from lenvatinib-resistant cells to sensitive cells and induce lenvatinib resistance of receipt cells.

Conclusion: CircPAK1 exerts its oncogenic function by competitively binding 14-3-3 ζ with YAP, thus promoting YAP nucleus localization, leading to the inactivation of a Hippo signaling pathway. Exosomal circPAK1 may drive resistance to lenvatinib, providing a potential therapeutic target for HCC patients.