PHB2 Promotes SHIP2 Ubiquitination via the E3 ligase NEDD4 to Regulate AKT Signaling in Gastric Cancer

Abstract

Background: Prohibitin 2 (PHB2) exhibits opposite functions of promoting or inhibiting tumour across various cancer types; nevertheless, its involvement in gastric cancer (GC) remains predominantly unexplored. Method: PHB2 protein expression levels in GC and normal tissues were examined using western blot and Immunohistochemistry. PHB2 expression level associations with patient outcomes were examined through Kaplan-Meier plotter analysis utilizing GEO datasets (GSE14210 and GSE29272). The biological role of PHB2 and its subsequent regulatory mechanisms were elucidated in vitro and in vivo. GC cell proliferation was assessed using MTT cell viability analysis, clonogenic assays, and BrdU incorporation assays, while the growth of GC xenografted tumors was measured via IHC staining of Ki-67. The interaction among PHB2 and SHIP2, as well as between SHIP2 and NEDD4, was identified through co-immunoprecipitation, GST pull-down assays, and deletion-mapping experiments. SHIP2 ubiquitination and degradation were assessed using Cycloheximide treatment, plasmid transfection and co-immunoprecipitation, followed by western blot analysis. Results: Our analysis revealed a substantial increase in PHB2 expression in GC tissues compared to adjacent normal tissues. Notably, higher PHB2 levels correlated with poorer patient outcomes, suggesting its clinical relevance. Functionally, silencing PHB2 in GC cells significantly reduced cell proliferation and retarded GC tumour growth, whereas overexpression of PHB2 further enhanced GC cell proliferation. Mechanistically, PHB2 physically interacted with Src homology 2-containing inositol 5-phosphatase 2 (SHIP2) in the cytoplasm of GC cells, thus leading to SHIP2 degradation via its novel E3 ligase NEDD4. It subsequently activated the PI3K/Akt signaling pathway and thus promoted GC cell proliferation. Conclusion: Our findings highlight the importance of PHB2 upregulation in driving GC progression and its association with adverse patient outcomes. Understanding the functional impact of PHB2 on GC growth contributes valuable insights into the molecular underpinnings of GC and may pave the way for the development of targeted therapies to improve patient outcomes.