Transfer RNA (tRNA)‐derived small RNAs (tsRNAs) have recently emerged as important regulators of protein translation and shown to have diverse biological functions. However, the underlying cellular and molecular mechanisms of tsRNA function in the context of dynamic cell‐state transitions remain unclear. Expression analysis of tsRNAs in distinct heterologous cell and tissue models of stem vs. differentiated states revealed a differentiation‐dependent enrichment of 5′‐tsRNAs. We report the identification of a set of 5′‐tsRNAs that is upregulated in differentiating mouse embryonic stem cells (mESCs). Notably, interactome studies with differentially enriched 5′‐tsRNAs revealed a switch in their association with “effector” RNPs and “target” mRNAs in different cell states. We demonstrate that specific 5′‐tsRNAs can preferentially interact with the RNA‐binding protein, Igf2bp1, in the RA‐induced differentiated state. This association influences the transcript stability and thereby translation of the pluripotency‐promoting factor, c‐Myc, thus providing a mechanistic basis for how 5′‐tsRNAs can modulate stem cell states in mESCs. Together our study highlights the role of 5′‐tsRNAs in defining distinct cell states.

The COVID-19 pandemic has strained testing capabilities worldwide. There is an urgent need to find economical and scalable ways to test more people. We present Tapestry, a novel quantitative nonadaptive pooling scheme to test many samples using only a few tests. The underlying molecular diagnostic test is any real-time RT-PCR diagnostic panel approved for the detection of the SARS-CoV-2 virus. In cases where most samples are negative for the virus, Tapestry accurately identifies the status of each individual sample with a single round of testing in fewer tests than simple two-round pooling. We also present a companion Android application BYOM Smart Testing which guides users through the pipetting steps required to perform the combinatorial pooling. The results of the pooled tests can be fed into the application to recover the status and estimated viral load for each individual sample. NOTE: This protocol has been validated with in vitro experiments that used synthetic RNA and DNA fragments and additionally, its expected behavior has been confirmed using computer simulations. Validation with clinical samples is ongoing. We are looking for clinical collaborators with access to patient samples. Please contact the corresponding author if you wish to validate this protocol on clinical samples.

Small non-coding RNAs such as miRNAs, piRNAs and endo-siRNAs fine-tune gene expression through post-transcriptional regulation, modulating important processes in development, differentiation, homeostasis and regeneration. Using deep sequencing, we have profiled small non-coding RNAs in Hydra magnipapillata and investigated changes in small RNA expression pattern during head regeneration. Our results reveal a unique repertoire of small RNAs in hydra. We have identified 126 miRNA loci; 123 of these miRNAs are unique to hydra. Less than 50% are conserved across two different strains of Hydra vulgaris tested in this study, indicating a highly diverse nature of hydra miRNAs in contrast to bilaterian miRNAs. We also identified siRNAs derived from precursors with perfect stem–loop structure and that arise from inverted repeats. piRNAs were the most abundant small RNAs in hydra, mapping to transposable elements, the annotated transcriptome and unique non-coding regions on the genome. piRNAs that map to transposable elements and the annotated transcriptome display a ping–pong signature. Further, we have identified several miRNAs and piRNAs whose expression is regulated during hydra head regeneration. Our study defines different classes of small RNAs in this cnidarian model system, which may play a role in orchestrating gene expression essential for hydra regeneration.