Cell-type-specific expression of tRNAs in the brain regulates cellular homeostasis
Defects in tRNA biogenesis are associated with multiple neurological disorders, yet our understanding of these diseases has been hampered an inability to determine tRNA expression in individual cell types within a complex tissue. Here, we developed a mouse model in which RNA Polymerase III is conditionally epitope-tagged in a Cre-dependent manner, allowing us to accurately profile tRNA expression in any cell type in vivo. We investigated tRNA expression in eleven diverse nervous system cell types, revealing dramatic heterogeneity in the expression of tRNA genes between populations. We found that while maintenance of levels of tRNA anticodon pools is critical for cellular homeostasis, neurons are differentially vulnerable to insults to distinct tRNA anticodon families. Cell type-specific translatome analysis suggests that the balance between tRNA availability and codon demand may underlie such differential resilience. Our work provides a platform for investigating the complexities of mRNA translation and tRNA biology in the brain. Overall design: The RiboTag mouse line (B6N.129-Rpl22tm1.1Psam/J mice) was crossed to appropriate Cre driver lines to obtain Rpl22 Fl/+, CRE + mice, expressing HA-tagged Rpl22 in the appropriate cell populations. The following populations were profiled: Cerebellar granule cells (Gabra6-cre, cerebellum), layer V cortical neurons (Rbp4-cre, cortex), dentate gyrus granule cells (Rbp4-cre, hippocamus), inhibitory interneurons (Dlx6a-cre, hippocampus), astrocytes (Aldh1-cre, cortex), and microglia (Tmem119-cre, cortex). HA-tagged RPL22 and associated mRNAs were immunoprecipitated using Pierce anti-HA magnetic beads (ThermoFisher Scientific) from the appropriate brain regions of 6-week-old mice, and mRNA libraries were made using the Kapa Stranded mRNA-Seq Kit (Roche) according to the manufacturer's protocol.The design includes three biological replicates for each cell type. For cerebellar granule cells, astrocytes, layer V cortical neurons, and dentate gyrus granule cells, one sample each was sequenced twice to account for batch effects.