Abstract Paralog genes are ubiquitous across the human genome, with nearly 70% of all human genes belonging to paralog gene families. Gene duplication throughout evolutionary history has provided genetic robustness, allowing human cells to survive if a single gene of an essential paralog pair is lost. Only recently have researchers been able to identify paralog genes required for cancer cell survival - through dual gRNA CRISPR screens, a subset of pan-essential paralogs has been identified across many cancer cell types. Interestingly, many groups have identified genetic compensation occurring between paralogous genes - in contexts where one paralog is lost through genetic means, expression of related genes increases. More recently, this phenomenon has been identified to be an active process, requiring degradation of mutant mRNA through the nonsense-mediated decay (NMD) pathway, and subsequent upregulation of sequence-similar genes. To assess how commonly this phenomenon occurs among essential paralog pairs, we developed a focused sgRNA library to generate single knockouts of 55 known essential paralog pairs. PC9 cells, a non-small cell lung cancer cell line, were transduced with this CRISPR library and subjected to perturb-seq to assess both gRNA and full single-cell transcriptome expression of cells following single paralog gene knockout. In this assay, we captured over 40,000 single cells with robust gRNA detection and full transcriptomic expression profiles, as well as target-enriched expression profiles of the 110 paralog genes of interest. Following differential gene expression analysis, six paralog pairs demonstrated measurable transcriptional adaptation, including the pan-essential paralogs TIA1/TIAL1 and NONO/PSPC1. Further validation of these pairs has confirmed that the TIA1/TIAL1 compensation occurs through degradation-dependent mechanisms; reduction in target gene expression through CRISPRi did not induce adapting gene expression, and pharmacological and genetic inhibition of the NMD-pathway was required. Further, nuclear import is an essential step in this adaptation process - blocking importin Β1 function through inhibitors or silencing of Importin 8 blocked this transcriptional adaptation. This work is the first to establish the prevalence of genetic compensation among essential paralogs in a high-throughput experimental model, and sheds light on this poorly described mechanism of degradation-dependent transcriptional adaptation. Citation Format: Siobhan O'Brien, Marissa Fujimoto, Amy Lowe, Alice Berger. Transcriptional adaptation among highly essential paralogs contributes to genetic robustness of cancer genomes abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 5916.
O'Brien et al. (Fri,) studied this question.