Lysosomal RNA profiling reveals targeting of specific types of RNAs for degradation

Autophagy targets a wide variety of substrates for degradation within lysosomes. Although lysosomes are known to possess RNase activity, the role of lysosomal RNA degradation in post-transcriptional gene regulation is not well understood. We defined RNASET2, PLD3, and both endogenous and exogenous RNase A family members as lysosomal RNases. Cells lacking these RNases accumulated large amounts of lysosomal RNA. Quantitative RNA sequencing of lysosomal contents revealed that although all RNA types can enter lysosomes, the SRP RNAs, Y RNAs, 5' TOP mRNAs, long-lived mRNAs, and mRNAs encoding membrane and secreted proteins were specifically enriched. This specificity was due to autophagy, and lysosomally degraded transcripts had autophagy-dependent changes in stability. To explore the basis of this specificity, we identified sequence motifs of SRP RNAs and 5' TOP mRNAs that are necessary and sufficient for enhanced lysosomal targeting. Our results establish lysosomes as selective modulators of cellular RNA content. Overall design: RNA sequencing of whole cell and lysosomal samples as well as RNA stability measurements using 5EU.