Pseudouridine guides small RNA transport and inheritance via Exportin-t

Pseudouridine (?) is an isomer of uridine found in ribosomal, transfer and other structural RNAs as well as in some mRNAs and non-coding RNAs, but is difficult to detect in short RNA sequences. Using modified techniques we found ? in microRNAs (miRNAs) and their precursors from mammalian and plant cells, primarily at the 5' terminus of the mature miRNA. Small RNAs targeting transposons in reproductive cells (piRNA in testis and easiRNA in pollen) were highly enriched for ?, indicating a potential role in epigenetic inheritance. In pollen, pseudouridylated small RNAs were produced by RNA polymerase IV and were localized to sperm cells, as were miRNAs with terminal ?. We show that pseudouridylated easiRNAs from pollen contribute to imprinting and the triploid block (chromosome dosage-dependent epigenetic lethality) via the activity of PAUSED/HEN5, the plant homolog of Exportin-t. Exportin-t is required for nuclear export of pseudouridylated tRNA, and we found that PSD is required for cell-cell transport of small RNA in the germline. Overall design: Small RNA-Seq experiment following RNA Immunoprecipation for pseudouridine (psi-IP) in WildType and Pus1 Knock Down 3T3 cells. Samples analysed include RNA following psi-IP as well as control Input samples. Time point analysed include 3 and 6 days after Knock Down of Pus1.