Endocrine Abstracts

Thyroid cancer progression is intricately linked to cell motility, a complex process that relies on the coordination of multiple signalling pathways, cell adhesion mechanisms, and actin dynamics. The proto-oncogene pituitary tumor-transforming gene (PTTG)-binding factor (PBF/PTTG1IP) significantly enhances thyroid cancer cell migration and invasion through Y174 phosphorylation by Src kinase. Phosphoproteomic and RNA-Seq analyses showed that PBF upregulation in Nthy-ori 3-1 thyroid cells altered expression and phosphorylation of adhesion and cytoskeletal proteins. We hypothesised that PBF is a physiological regulator of cell adhesion, and oncogenic expression promotes motility by altering adhesion dynamics. CRISPR/Cas9-mediated PBF-knockout (KO) TPC-1 papillary thyroid carcinoma cells exhibited significantly decreased adhesion on fibronectin compared with control TPC-1 cells. Immunofluorescence staining revealed altered structure and distribution of focal adhesions (FAs), which connect the cytoskeleton and extracellular matrix. PBF-depleted TPC-1 cells had markedly reduced focal adhesion kinase (FAK), vinculin and paxillin staining with fewer, shorter FAs predominantly around the cell periphery, whereas control cells displayed numerous, elongated FAs along actin fibers. Pbf-KO mouse embryonic fibroblasts (MEFs) also demonstrated decreased cell-substrate adhesion and altered FAs. Early adhesion assays (15-30 min) demonstrated reduced phosphorylated FAK-Y397 and fewer FAK-associated early FA structures at the Pbf-KO MEF peripheral membrane, while wild-type MEFs formed early FAK/paxillin adhesions that gradually matured over time. Live-cell LifeAct-GFP imaging indicated impaired spreading, lamellipodia formation and lack of directionality in PBF-KO TPC-1 and MEF cells. During migration, Pbf-KO MEF cells displayed disorganised Golgi lacking orientation towards the leading edge, suggesting impaired polarity. These findings provide important insights into FA dynamics in PBF-induced thyroid cancer cell motility, revealing a critical role for PBF in regulating early adhesion assembly. Further investigations are required to elucidate the precise interactions of PBF with cell adhesion protein complexes, which may ultimately reveal new therapeutic targets in thyroid cancer progression.