circE2F1-encoded peptide inhibits circadian machinery essential for nucleotide biosynthesis and tumor progression via repressing SPIB/E2F1 axis

Circadian clock dominates a variety of biological activities, while its roles and regulatory mechanisms in neuroblastoma (NB), a pediatric extracranial malignancy, still remain largely elusive. Herein, through comprehensive analyses of public datasets, E2F transcription factor 1 (E2F1) and its circular RNA (circE2F1)-encoded 99-amino acid peptide (E2F1-99aa) were identified as vital regulators of circadian machinery essential for purine and pyrimidine biosynthesis during NB progression. Mechanistically, through interaction with Spi-B transcription factor (SPIB), E2F1 was transactivated to up-regulate circadian machinery genes (CRY1 and TIMELESS), resulting in relief of CLOCK/BMAL1-repressed transcription of enzymes (DHODH, PAICS, or PPAT) essential for de novo purine and pyrimidine biosynthesis. The biogenesis of circE2F1 was repressed by eukaryotic translation initiation factor 4A3 (EIF4A3), while E2F1-99aa or its truncated peptide competitively bound to SPIB, leading to decrease in SPIB-E2F1 interaction, circadian machinery and nucleotide biosynthetic gene expression, purine or pyrimidine biosynthesis, tumorigenesis, and aggresiveness of NB cells. In clinical NB cases, high EIF4A3, E2F1 or SPIB levels were correlated with low survival possibility of patients, while lower circE2F1 or E2F1-99aa levels were associated with advanced stages and tumor progression. These results indicate that circE2F1-encoded peptide inhibits circadian machinery essential for nucleotide biosynthesis and tumor progression via repressing SPIB/E2F1 axis.