A RiboCancer cell line panel reveals that CLL‐associated Rps15 mutations translationally rewire transcription through codon‐specific tRNA accommodation defects

Abstract Recurrent point mutations in ribosomal proteins (RPs) RPL10 and RPS15 are found in T‐cell acute lymphoblastic leukemia (T‐ALL) and chronic lymphocytic leukemia (CLL), respectively. Furthermore, deletions of RPL5, RPL11, and RPL22 are frequent in hematologic diseases such as Diamond Blackfan Anemia, T‐ALL, multiple myeloma, and in a variety of solid tumors. Yet, the role of these RP defects in dysregulation of the ribosomal translation function remains poorly understood. We engineered an isogenic RiboCancer cell line library modeling the most recurrent RP defects in blood and solid cancers and characterized it by a multi‐omics translatome analysis (proteome, Ribo‐seq, and total RNA‐seq) as well as RiboMethSeq. Within this RiboCancer panel, CLL ‐ associated Rps15 mutations induced the strongest alterations in mRNA translation, affecting up to 10% of expressed genes. Cryo‐electron microscopy revealed that these mutations destabilize the Rps15 C‐terminus and affect the translation elongation cycle dynamics by deregulating accommodation of aminoacylated tRNAs at the ribosomal A‐site. This accommodation defect showed specificity for 11 codons, explaining the reduced translation efficiency of genes with high presence of these codons in Rps15‐mutant cells. Notably, these genes were enriched for epigenetic and transcriptional regulators such as transcription factor Runx3, resulting in downregulation of Runx3 target genes involved in immune regulation. By developing and characterizing a unique RiboCancer cell line panel, we mapped translational rewiring driven by the most frequent somatic RP mutations. We provide unprecedented mechanistic insights into translation defects induced by CLL‐associated Rps15 mutations, and reveal an intriguing translation‐based rewiring of transcription in CLL.