The adaptive architecture of tRNA dependencies across physiological tumor environments

The genetic code is decoded by a highly redundant transfer RNA (tRNA) repertoire, yet whether this apparent redundancy serves a functional role beyond ensuring robust translation remains unclear. Here, we establish an atlas of tRNA dependencies through large-scale CRISPRi perturbation mapping across matched in vivo tumor growth and in vitro cell culture for 16 cancer cell lines spanning seven tissue types. During in vivo tumor growth, tRNA dependencies exhibited tissue organization at isodecoder resolution and were associated with codon demand at the isoacceptor level. Strikingly, these relationships were reorganized when cells were cultured in vitro, establishing environmental sensitivity of tRNA dependency--in contrast to protein components of the decoding machinery such as aminoacyl-tRNA synthetases (aaRSs), whose dependencies were largely preserved across environments. Despite pronounced context specificity, environmental alterations in tRNA dependency were dominated by a shared response across cancer cell lines, organized across isoacceptor and isodecoder levels. By disentangling stable cellular identity from environment-associated dependency states, we resolved a shared axis of isoacceptor dependency remodeling and identified specific tRNA isoacceptors associated with promoting or restraining adaptation to the tumor environment. We further connect this functional architecture to differential translation of codon-usage-biased programs associated with proliferation and tumor-microenvironmental stress. Together, our study reveals that tRNA redundancy does not imply functional equivalence, but instead forms a structured and environmentally plastic control layer linking coding information to translational output.