26Background: Diverse biological processes and transcriptional programs are 27 regulated by RNA polymerase II (Pol II), which is recruited by the general transcription 28 machinery to the core promoter to initiate transcription. TRF2 (TATA-box-binding 29 protein-related factor 2) is an evolutionarily conserved general transcription factor that 30 is essential for embryonic development of Drosophila melanogaster, C. elegans, 31 zebrafish and Xenopus. Nevertheless, the cellular processes that are regulated by 32 TRF2 are largely underexplored. 33Results: Here, using Drosophila Schneider cells as a model, we discovered that TRF2 34 regulates apoptosis and cell cycle progression. We show that TRF2 knockdown 35 results in increased expression of distinct pro-apoptotic genes and induces apoptosis. 36Using flow cytometry, high-throughput microscopy and advanced imaging-flow 37 cytometry, we demonstrate that TRF2 regulates cell cycle progression and exerts 38 distinct effects on G1 and specific mitotic phases. RNA-seq analysis revealed that 39 TRF2 controls the expression of Cyclin E and the mitotic cyclins, Cyclin A, Cyclin B 40 and Cyclin B3, but not Cyclin D or Cyclin C. To identify proteins that could account for 41 the observed regulation of these cyclin genes, we searched for TRF2-interacting 42 proteins. Interestingly, mass spectrometry analysis of TRF2-containing complexes 43 identified GFZF, a nuclear glutathione S-transferase implicated in cell cycle regulation, 44and Motif 1 binding protein (M1BP). TRF2 has previously been shown to interact with 45 M1BP and M1BP has been shown to interact with GFZF. Furthermore, available ChIP-46 exo data revealed that TRF2, GFZF and M1BP co-occupy the promoters of TRF2-47 with TRF2, it is TRF2, rather than GFZF or M1BP, that is the main factor regulating 50 the expression of Cyclin E and the mitotic cyclins. 51 Conclusions: Our findings uncover a critical and unanticipated role of a general 52 transcription factor as a key regulator of cell cycle and apoptosis. 53 54 Keywords 55 Basal transcription machinery, RNA polymerase II, gene expression, TATA box-56 binding protein (TBP), TBP-related factor 2 (TRF2), cyclin genes. 57 58 BACKGROUND 59 Multiple biological processes and transcriptional programs are regulated by RNA 60 polymerase II (Pol II). The initiation of transcription of protein-coding genes and 61 distinct non-coding RNAs occurs following the recruitment of Pol II to the core 62 promoter region by the general/basal transcription machinery (1-4). The core 63promoter, which directs accurate initiation of transcription and encompasses the 64 transcription start site (TSS), may contain short DNA sequence elements/motifs, 65 which confer specific properties to the core promoter (1, 4-10). The first step in the 66 recruitment of Pol II to initiate transcription is the binding of TFIID, which is 67 composed of TATA-box-binding protein (TBP) and TBP-associated factors. 68Remarkably, although TBP is considered a universal general transcription factor, 69 robust Pol II transc...