Functional CRISPR screen identifies AP1-associated enhancer regulating FOXF1 to modulate oncogene-induced senescence

Abstract

BackgroundFunctional characterization of non-coding elements in the human genome is a major genomic challenge and the maturation of genome-editing technologies is revolutionizing our ability to achieve this task. Oncogene-induced senescence, a cellular state of irreversible proliferation arrest that is enforced following excessive oncogenic activity, is a major barrier against cancer transformation; therefore, bypassing oncogene-induced senescence is a critical step in tumorigenesis. Here, we aim at further identification of enhancer elements that are required for the establishment of this state.ResultsWe first apply genome-wide profiling of enhancer-RNAs (eRNAs) to systematically identify enhancers that are activated upon oncogenic stress. DNA motif analysis of these enhancers indicates AP-1 as a major regulator of the transcriptional program induced by oncogene-induced senescence. We thus constructed a CRISPR-Cas9 sgRNA library designed to target senescence-induced enhancers that are putatively regulated by AP-1 and used it in a functional screen. We identify a critical enhancer that we name EnhAP1-OIS1 and validate that mutating the AP-1 binding site within this element results in oncogene-induced senescence bypass. Furthermore, we identify FOXF1 as the gene regulated by this enhancer and demonstrate that FOXF1 mediates EnhAP1-OIS1 effect on the senescence phenotype.ConclusionsOur study elucidates a novel cascade mediated by AP-1 and FOXF1 that regulates oncogene-induced senescence and further demonstrates the power of CRISPR-based functional genomic screens in deciphering the function of non-coding regulatory elements in the genome.Electronic supplementary materialThe online version of this article (10.1186/s13059-018-1494-1) contains supplementary material, which is available to authorized users.

Defining a Characteristic Gene Expression Set Responsible for Cancer Stem Cell-Like Features in a Sub-Population of Ewing Sarcoma Cells CADO-ES1

“…AP-1 is also showing up in Kaposi’s sarcoma-associated herpesvirus [60] and adult T-cell leukemia [61,62,63]. A nice AP-1-associated enhancer regulation motif based on FOXF1 [64] might also be a model for the family member FOXO1, which is exposed in our study and is associated with Rhabdomyosarcoma. A publication of Lopez-Bergami et al [65] gives a broader overview on AP-1 pathways in cancer and has a nice crosslink in his Table 2 ‘ATF2 transcriptional targets’, the cell cycle-associated cyclin CCND1.…”

A lncRNA-SWI/SNF complex crosstalk controls transcriptional activation at specific promoter regions

“…factor already described as involved in SWI/SNF complex recruitment 33 and in enhancer regulation upon senescence induction 34 . These data suggest that SMARCB1 ChIP-seq is mapping SWI/SNF binding loci.…”

Beyond the Exome: The Non-coding Genome and Enhancers in Neurodevelopmental Disorders and Malformations of Cortical Development

“…Using a similar approach, more than 18.000 gRNAs were used to test around 700 kb of sequence flanking genes involved in BRAF inhibitor resistance in melanoma, finding non-coding regions involved in gene regulation and chemotherapeutic resistance (Sanjana et al, 2016). Other studies investigated putative enhancers involved in oncogene induced senescence (Han et al, 2018), regulation of the HPRT gene involved in Lesch–Nyhan syndrome (Gasperini et al, 2017) and regulation of the POU5F1 gene in embryonic stem cells (Diao et al, 2016, 2017), amongst others (Canver et al, 2015, 2017; Rajagopal et al, 2016; Sen et al, 2016). Besides genome-engineering, CRISPR-Cas9 can also be applied to edit the epigenome, and also this can be coupled to high-throughput screening.…”