Oncogenic effect of PHLDB2 is associated with epithelial-mesenchymal transition and E-cadherin regulation in colorectal cancer

Abstract

Background: Pleckstrin Homology Like Domain Family Member 2 (PHLDB2) is an important protein with a PH-domain for interaction with partners to regulate cell migration. However, the role of PHLDB2 in human cancer metastasis, especially in colon cancer, still remains elusive. Methods: The RNA-seq and clinical data of colorectal cancer patients from the Cancer Genome Atlas (TCGA) were analyzed for correlations between PHLDB2 and clinical outcomes as well as epithelial-mesenchymal transition (EMT) markers. Wound healing and transwell invasion assays were used to determine the effects of PHLDB2 on cell migration and invasiveness. Western blot and qRT-PCR analyses were employed to detect protein and mRNA changes, respectively. Co-immunoprecipitation was performed to assess protein-protein interaction. Results: In the present report, by following our previous study, we found that PHLDB2 expression is associated with poorer prognosis, including disease-free survival, tumor stage, nodes pathology, as well as lymphatic and vascular invasion through TCGA data analysis. In addition, PHLDB2 expression is highly correlated with multiple epithelial-mesenchymal transition (EMT) markers involving cell-surface proteins (N-cadherin and OB-cadherin), cytoskeletal markers (α-SMA and Vimentin), ECM proteins (Fibronectin and Laminin 5), and transcription factors (Snail2, ZEB1, and Ets-1). We also demonstrated that PHLDB2 knockdown mediated by siRNA was sufficient to attenuate colon cancer cell migration and invasion, as well as E-Cadherin reduction, by TGF-β treatment. Interestingly, PHLDB2 expression levels were significantly elevated in response to EMT induction by TGF-β and EGF. Moreover, we found that PHLDB2 could bind to MDM2 and facilitate MDM2-mediated E-Cadherin degradation. Conclusions: Our findings suggest that PHLDB2 is a downstream effector of EMT pathway and may present as an important biomarker for colon cancer prognosis and a target for colon cancer intervention.