TNIK Regulates Cytoskeletal Organization to Promote Focal Adhesion Turnover and Mitosis in Lung Adenocarcinoma.

Abstract

Background: Lung cancer is the primary cause of cancer-related mortality, but the molecular mechanisms behind this malignancy remain unclear.

Methods: The Cancer Genome Atlas (TCGA) online database and tissue chips were used to analyze the expression levels of tumor necrosis factor receptor-associated factor 2 (TRAF2)- and non-catalytic region of tyrosine kinase adaptor protein (NCK)- interacting kinase (TNIK) protein in lung cancer. A549 and PC-9 lung adenocarcinoma (LUAD) cells with stable TNIK knockdown were generated by lentivirus infection. The tumor phenotypes were subsequently examined both in vitro and in vivo. The TCGA online database and RNA-sequencing of TNIK-knockdown cells were used to study the molecular mechanism underlying the TNIK-mediated phenotype of LUAD cells. The effects of TNIK knockdown on focal adhesion dynamics and mitosis were examined by indirect immunofluorescence and Western blot, on the sensitivity to chemotherapy drugs by cell counting kit-8 (CCK-8) assay, on apoptosis by flow cytometry, and on cell proliferation by 5-ethynyl-2'-deoxyuridine (EDU).

Results: TNIK was highly expressed in LUAD (p < 0.0001), predominantly in the cytosol. Phenotype assays revealed that TNIK knockdown in LUAD cells led to a significant increase in cell spreading (p < 0.0001), but also inhibition of cell growth and movement (p < 0.01). Mechanistically, TNIK was found to regulate F-actin and microtubule organization, as well as the Ras homolog gene family (RHO)/RHO-associated kinase 2 (ROCK2)/LIM motif-containing protein kinase 1 (LIMK1) signaling pathway, thereby playing a crucial role in the control of focal adhesion turnover and mitosis. Additionally, the silencing of TNIK enhanced the sensitivity of LUAD cells to chemotherapeutic drugs.

Conclusions: Our findings suggest that TNIK regulates focal adhesion turnover and mitosis to promote tumor malignancy via the RHO/ROCK2/LIMK1 pathway. The combination of TNIK targeting with chemotherapeutic drugs could be an effective strategy to overcome resistance in LUAD.