ARPC1B promotes mesenchymal phenotype maintenance and radiotherapy resistance by blocking TRIM21-mediated degradation of IFI16 and HuR in glioma stem cells

Intratumoral heterogeneity is the primary challenge in the treatment of glioblastoma (GBM). The presence of glioma stem cells (GSCs) and their conversion between different molecular phenotypes contribute to the complexity of heterogeneity, culminating in preferential resistance to radiotherapy. ARP2/3 (actin-related protein-2/3) complexes (ARPs) are associated with cancer migration, invasion and differentiation, while the implications of ARPs in the phenotype and resistance to radiotherapy of GSCs remain unclear.

ARPC1B promoted MES phenotype maintenance and radiotherapy resistance by inhibiting TRIM21-mediated degradation of IFI16 and HuR, thereby activating the NF-κB and STAT3 signaling pathways, respectively. AZD6738, identified based on ARPC1B expression, exhibited excellent anti-GSC activity in combination with radiotherapy.

We screened the expression of ARPs in TCGA-GBM and CGGA-GBM databases. Tumor sphere formation assays and limiting dilution assays were applied to assess the implications of ARPC1B in tumorigenesis. Apoptosis, comet, γ-H2AX immunofluorescence (IF), and cell cycle distribution assays were used to evaluate the effect of ARPC1B on radiotherapy resistance. Immunoprecipitation (IP) and mass spectrometry analysis were used to detect ARPC1B-interacting proteins. Immune blot assays were performed to evaluate protein ubiquitination, and deletion mutant constructs were designed to determine the binding sites of protein interactions. The Spearman correlation algorithm was performed to screen for drugs that indicated cell sensitivity by the expression of ARPC1B. An intracranial xenograft GSC mouse model was used to investigate the role of ARPC1B in vivo.

We concluded that ARPC1B was significantly upregulated in MES-GBM/GSCs and was correlated with a poor prognosis. Both in vitro and in vivo assays indicated that knockdown of ARPC1B in MES-GSCs reduced tumorigenicity and resistance to IR treatment, whereas overexpression of ARPC1B in PN-GSCs exhibited the opposite effects. Mechanistically, ARPC1B interacted with IFI16 and HuR to maintain protein stability. In detail, the Pyrin of IFI16 and RRM2 of HuR were implicated in binding to ARPC1B, which counteracted TRIM21-mediated degradation of ubiquitination to IFI16 and HuR. Additionally, the function of ARPC1B was dependent on IFI16-induced activation of NF-κB pathway and HuR-induced activation of STAT3 pathway. Finally, we screened AZD6738, an ataxia telangiectasia mutated and rad3-related (ATR) inhibitor, based on the expression of ARPC1B. In addition to ARPC1B expression reflecting cellular sensitivity to AZD6738, the combination of AZD6738 and radiotherapy exhibited potent antitumor effects both in vitro and in vivo.