CRISPRa screens reveal RNA sensitisation routes

In melanoma cells engineered with CRISPRa (dCas9-VPR), a large pooled screen was performed under pressure from TCR-specific CD8 T cells. This revealed a diverse set of genes whose activation either protects or sensitises cells to cytotoxicity, spanning transcription factors, chromatin regulators and RNA-processing proteins. Many sensitising effects were independent of baseline cell fitness, pointing to a form of 'immune RNA-based synthetic lethality', where gene expression alone is not harmful but becomes detrimental in the presence of T cells. CASP3 illustrates this principle: its overexpression leaves cells viable in isolation but primes them for apoptosis once T cells engage, with activation of caspase-3 occurring specifically during immune attack.

Follow-up Perturb-seq experiments place these genes within shared regulatory networks, showing that distinct perturbations converge on common pathways involving apoptosis, cytokine signalling and cell–cell communication. Notably, Wnt ligands emerged as mediators that can directly enhance T cell activity, highlighting bidirectional interactions between tumour and immune cells. Validation in vivo and with spatially resolved in situ Perturb-seq confirms that regulators such as MYC and SAFB can reproducibly sensitise tumour cells, with SAFB acting in a particularly TCR-specific manner without broadly altering cell state. Together, the work demonstrates how CRISPRa can systematically reveal RNA-driven mechanisms that selectively restore immune-mediated elimination of cancer cells.

The study was led by Reece Villarin Akana, Jeehyun Yoe and Livnat Jerby at Stanford University. It was published in Nature Genetics on 8 April 2026.