Rolling-translated EGFR variants sustain EGFR signaling and promote glioblastoma tumorigenicity

Background. Aberrant epidermal growth factor receptor (EGFR) activation is observed in over 50% of cases of adult glioblastoma (GBM). Nevertheless, EGFR antibodies are ineffective in clinical GBM treatment, suggesting the existence of redundant EGFR activation mechanisms. Whether circular RNA (circRNA) encodes a protein involved in EGFR-driven GBM remains unclear. We reported an unexpected mechanism in which circular EGFR RNA (circEGFR) encodes a novel EGFR variant to sustained EGFR activation. Method. We used RNA-seq, Northern blot, and Sanger sequencing to confirm the existence of circ-EGFR. Antibodies and a liquid chromatograph tandem mass spectrometer were used to identify circ-EGFR protein products. Lentivirus-transfected stable cell lines were used to assess the biological functions of the novel protein in vitro and in vivo. Clinical implications of circ-EGFR were assessed using 97 pathologically diagnosed GBM patient samples. Results. The infinite open reading frame (iORF) in circ-EGFR translated repeating amino acid sequences via rolling translation and programmed -1 ribosomal frameshifting (-1PRF) induced out-of-frame stop codon (OSC), forming a polymetric novel protein-complex, which we termed rolling-translated EGFR (rtEGFR). rtEGFR directly interacted with EGFR, maintained EGFR membrane localization and attenuated EGFR endocytosis and degradation. Importantly, circ-EGFR levels correlated with the EGFR signature and predicted the poor prognosis of GBM patients. Deprivation of rtEGFR in brain tumor-initiating cells (BTICs) attenuated tumorigenicity and enhanced the anti-GBM effect. Conclusion. Our findings identified the endogenous rolling-translated protein and provided strong clinical evidence that targeting rtEGFR could improve the efficiency of EGFR-targeting therapies in GBM.

Automated summary

The study identified a novel EGFR variant, rtEGFR, encoded by circEGFR, which promotes GBM development by maintaining EGFR activation and inhibiting EGFR degradation, showing significant correlation with patient prognosis. Targeting rtEGFR could potentially enhance the efficacy of EGFR-targeting therapies in GBM treatment.