Understanding gene regulatory networks is fundamental to molecular biology. Transcription factors are critical components of these networks, as they bind to specific genomic sequences to regulate gene expression and influence cell fate decisions. TFs regulate diverse physiological pathways, including immune responses, yet their systematic roles in tumour immune evasion remain poorly defined.
To systematically investigate this, we leveraged the Multiplexed Overexpression of Regulatory Factors (MORF) library — a barcoded open reading frame (ORF) collection encompassing over 3,500 human transcription factor isoforms — to perform a comprehensive functional screen in melanoma cells. This approach enabled high-throughput overexpression of individual TFs to assess their impact on tumour susceptibility to T cell–mediated killing.
Through this large-scale functional screen, we identified several known regulators of immune evasion, as well as previously uncharacterized transcription factors that confer resistance to cytotoxic T cells. Follow-up validation experiments, including targeted overexpression and co-culture assays, confirmed their phenotypic effects and robustness of the hits.
Building on these discoveries, we are now applying RNA-seq and ChIP-seq analyses to delineate the downstream gene networks and chromatin interactions driven by these candidate TFs. Together, our work provides a comprehensive functional map of transcription factors driving immune evasion in solid tumors and establishes a foundation for uncovering the molecular mechanisms underlying immunotherapy resistance.