AXL Inactivation Inhibits Mesothelioma Growth and Migration via Regulation of p53 Expression.

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Simple SummaryPrevious studies have shown that AXL is a crucial protein that is activated in mesothelioma to cause growth and invasiveness. p53 is the tumor suppressor protein that is most frequently inactivated by mutations in human cancers, but these p53 inactivating mutations are not common in mesothelioma. In the studies reported herein we demonstrate that AXL activation causes p53 functional inactivation in mesothelioma. Specifically, we show that AXL suppresses p53 expression by binding to DNA sequences upstream from the p53 gene, thereby blocking transcription of p53 DNA into RNA. We also show that AXL inhibition by a selective drug inhibits mesothelioma cell viability, migration, and invasion. p53 inactivation attenuates the impact of the AXL inhibitor, providing further support for interplay between AXL and p53 in mesothelioma oncogenesis. These studies demonstrate a novel feedback regulation loop between AXL and p53, and provide a rationale for mesothelioma therapies targeting AXL/p53 signaling.Malignant mesothelioma is a locally aggressive and highly lethal neoplasm. Dysregulation and activation of Gas6/AXL tyrosine kinase signaling are associated with mesothelioma progression, but the mechanisms of these AXL tumorigenic roles are poorly understood. p53 mutants in lung carcinoma upregulate AXL expression by binding and acetylating the AXL promoter. Although TP53 mutations are uncommon in mesothelioma, we hypothesized that these tumors might have alternative feedback mechanisms between AXL and p53. In the current report, we investigated AXL regulation of TP53 transcription, expression, and biological function in mesothelioma. AXL expression was stronger in mesothelioma than most of the other tumor types from the TCGA gene expression profile dataset. AXL knockdown by shRNA induced wild-type and mutant p53 expression in mesothelioma cell lines, suggesting that AXL pro-tumorigenic roles result in part from the suppression of p53 function. Likewise, induced AXL inhibited expression of wild type p53 in COS-7 cells and 293T cells. Immunofluorescence staining showed nuclear colocalization of AXL and p53; however, association of AXL and p53 was not demonstrated in immunoprecipitation complexes. The AXL effects on p53 expression resulted from the inhibition of TP53 transcription, as demonstrated by qRT-PCR after AXL silencing and TP53 promotor dual luciferase activity assays. Chromatin immunoprecipitation-qPCR and sequencing showed that AXL bound to the initial 600 bp sequence at the 5′ end of the TP53 promoter. AXL inhibition (shRNA or R428) reduced mesothelioma cell viability, migration, and invasion, whereas TP53 shRNA knockdown attenuated antiproliferative, migration, and invasive effects of AXL silencing or AXL inactivation in these cells. These studies demonstrate a novel feedback regulation loop between AXL and p53, and provide a rationale for mesothelioma therapies targeting AXL/p53 signaling.