Actin-like 6A predicts poor prognosis of hepatocellular carcinoma and promotes metastasis and epithelial-mesenchymal transition.
Bottom Line: Opposite results are observed when ACTL6A is knocked down.ACTL6A knockdown has the equal blockage effect as the Notch signaling inhibitor, N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butylester, in HCC cells.Further studies indicate that ACTL6A might manipulate SRY (sex determining region Y)-box 2 (SOX2) expression and then activate Notch1 signaling.
Affiliation: Liver Cancer Laboratory, Department of Surgery, Xiangya Hospital, Central South University, Changsha, China.Show MeSH
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Mentions: In this study, PLC/PRF5‐ACTL6A cells exhibited spindle‐like mesenchymal morphology, whereas PLC/RPF5 cells mostly looked like epithelial cobblestones in appearance. On the contrary, Hep3B‐shACTL6A cells changed to an epithelial phenotype as compared with mesenchymal‐like Hep3B cells (Fig. 3A). Considering the morphological change and the function of ACTL6A in HCC, we speculated that an abnormal high level of ACTL6A would trigger EMT.15, 17 This speculation was confirmed by cytological and histology experiments. Immunofluorescence (IF) analysis showed that PLC/PRF5‐ACTL6A cells presented an elongated cellular morphology and the appearance of F‐actin fibers compared with PLC/PRF5 cells. Inversely, Hep3B‐shACTL6A cells exhibited a cobblestone shape and shrinkable F‐actin fiber changes to Hep3B cells (Fig. 3B). The anchorage‐independent growth assay indicated that colonies that grew from PLC/PRF5‐ACTL6A cells were larger and more in soft agar than PLC/PRF5 cells, whereas colonies that grew from Hep3B‐shACTL6A cells were smaller and less than Hep3B cells (Fig. 3C). IF analysis further showed that ectopic expression of ACTL6A reduced expression of the epithelial marker, E‐cadherin, and increased the mesenchymal marker, vimentin, in PLC/PRF5‐ACTL6A cells, whereas knockdown ACTL6A in Hep3B cells increased E‐cadherin expression and decreased vimentin expression (Fig. 3D). Consistent with this, ACTL6A highly expressed cells showed decreased E‐cadherin and increased vimentin and snail (EMT transcriptional factor) expressions at both protein and mRNA levels, whereas ACTL6A knockdown induced the opposite results (Fig. 3E and Supporting Fig. 6). Moreover, IHC for HCC consecutive sections revealed that high ACTL6A expression was colocation with low E‐cadherin and high vimentin expression, and vice versa in ANLT (Fig. 3F). The correlation analysis revealed that ACTL6A expression was positively correlative with vimentin and negatively correlative with E‐cadherin expression in HCC samples in both the training and validation cohorts (Fig. 3F). These results indicated that ACTL6A played an important role in promoting EMT in HCC.
Affiliation: Liver Cancer Laboratory, Department of Surgery, Xiangya Hospital, Central South University, Changsha, China.