Expression of AE1/p16 promoted degradation of AE2 in gastric cancer cells.

BACKGROUND: Human anion exchanger 1 and 2 (AE1 and AE2) mediate the exchange of Cl(-)/HCO3 (-) across the plasma membrane and regulate intracellular pH (pHi). AE1 is specifically expressed on the surface of erythrocytes, while AE2 is widely expressed in most tissues, and is particularly abundant in parietal cells. Previous studies showed that an interaction between AE1 and p16 is a key event in gastric cancer (GC) progression, but the importance of AE2 in GC is unclear. METHODS: The relationship among AE1, AE2 and p16 in GC cells was characterized by molecular and cellular experiments. AE2 expression and pHi were measured after knockdown or forced expression of AE1 or p16 in GC cells. The effect of AE2 on GC growth and the correlation of AE2 expression with differentiation and prognosis of GC were also evaluated. The effect of gastrin on AE2 expression and GC growth was investigated in cellular experiments and mouse xenograft models. RESULTS: p16 binds to both AE1 and AE2 simultaneously. AE1 or p16 silencing elevated AE2 expression on the plasma membrane where it plays a role in pHi regulation and GC suppression. AE2 expression was decreased in GC tissue, and these decreased levels were correlated with poor differentiation and prognosis of GC. The low AE2 protein levels are due to rapid ubiquitin-mediated degradation that was facilitated in the presence of p16. Gastrin inhibited the growth of GC cells at least partially through up-regulation of AE2 expression. CONCLUSION: AE1/p16 expression promoted AE2 degradation in GC cells. Gastrin is a potential candidate drug for targeted therapies for AE1- and p16-positive GC.

PCAF acts as a gastric cancer suppressor through a novel PCAF-p16-CDK4 axis.

Gastric cancer (GC) is a leading cause of cancer-related death worldwide and the pathogenesis of GC remains largely unknown. Here, we demonstrate a novel mechanism by which P300/CBP associating factor (PCAF) acts as a tumor suppressor in GC cells. We showed that both PCAF mRNA and protein were downregulated in GC cells, and that this downregulation correlated with poor survival. Meanwhile, the interaction between human anion exchanger 1 (AE1) and p16 is a key event in GC development. We found that PCAF inhibited GC growth by interacting with AE1 and p16 to promote ubiquitin-mediated degradation of AE1 and p16 upregulation and translocation into the nucleus. Binding of nuclear p16 to CDK4 prevented the CDK4-Cyclin D1 interaction to inhibit GC proliferation. Furthermore, reduced PCAF levels in GC cells were associated with intracellular alkalinization and decreased immunity. Together these results suggest that PCAF acts as a GC suppressor through a novel PCAF-p16-CDK4 axis. The downregulation of PCAF expression in GC cells that follows intracellular alkalinization and decreased immune response, indicates that GC therapies should focus on restoring PCAF levels.

Direct interaction and cooperative role of tumor suppressor p16 with band 3 (AE1).

By using the C-terminal 112-residue of band 3 to screen the K562 cDNA library, we find that the p16 interacts with band 3, which was confirmed both in yeast and in mammalian cells. Functional experiments show that p16 facilitates the movement of band 3 to plasma membrane with increased anion transport activity in 293t cells. Moreover, expression of endogenous p16 in 293t cells was increased at 24 and 36 h after transfection with band 3. Our findings provide a novel regulation pathway for both band 3 and p16.