Author Contributions S.J. and C.L.K. designed and coordinated analysis of single-cell data. S.J. and A.B.-C. performed the majority of the scRNA-seq analyses and visualizations. J.M. and G.B. contributed to the CNA analysis. Y.H. contributed to the algorithm for marker gene discovery. F.M.G.C., M.C., A.B.-C. and S.J. analyzed transcription factor activity in the scRNA-seq data. N.D.J., S.H. and S.J. contributed to the analysis of the bulk RNA-seq data and the data availability submission. M.V. contributed to timed mating and tissue isolation in developing mouse embryos. D.F., M.V. and L.K.D. and contributed to primary tissue isolation, preparation and production of scRNA-seq libraries. B.K. performed all experiments in cellular models. L.G., S.J., W.T.F. and K.K.M. contributed to literature review and cell cluster annotations. L.G. provided expert advice on identification of developing pre-cerebellar populations. M.K.M. and L.G.M. contributed to the clinical annotation of tumor samples. P.-E.L. and G.T. provided bulk adult human brain RNA-seq samples. M.R., B.P. and A.A. provided human fetal brain samples.

The eukaryotic translation initiation factor eIF4E acts in the nuclear export and translation of a subset of mRNAs. Both of these functions contribute to its oncogenic potential. While the biochemical mechanisms that underlie translation are relatively well understood, the molecular basis for eIF4E's role in mRNA export remains largely unexplored. To date, over 3000 transcripts, many encoding oncoproteins, were identified as potential nuclear eIF4E export targets. These target RNAs typically contain a ∼50-nucleotide eIF4E sensitivity element (4ESE) in the 3 ′ UTR and a 7-methylguanosine cap on the 5 ′ end. While eIF4E associates with the cap, an unknown factor recognizes the 4ESE element. We previously identified cofactors that functionally interacted with eIF4E in mammalian cell nuclei including the leucine-rich pentatricopeptide repeat protein LRPPRC and the export receptor CRM1/XPO1. LRPPRC simultaneously interacts with both eIF4E bound to the 5 ′ mRNA cap and the 4ESE element in the 3 ′ UTR. In this way, LRPPRC serves as a specificity factor to recruit 4ESE-containing RNAs within the nucleus. Further, we show that CRM1 directly binds LRPPRC likely acting as the export receptor for the LRPPRC-eIF4E-4ESE RNA complex. We also found that Importin 8, the nuclear importer for cap-free eIF4E, imports RNA-free LRPPRC, potentially providing both coordinated nuclear recycling of the export machinery and an important surveillance mechanism to prevent futile export cycles. Our studies provide the first biochemical framework for the eIF4E-dependent mRNA export pathway.

Extracellular vesicles (EVs) are membrane-enclosed nanoparticles containing specific repertoires of genetic material. In mammals, EVs can mediate the horizontal transfer of various cargos and signaling molecules, notably miRNA and mRNA species. Whether this form of intercellular communication prevails in other metazoans remains unclear. Here, we report the first parallel comparative morphologic and transcriptomic characterization of EVs from Drosophila and human cellular models. Electronic microscopy revealed that human and Drosophila cells release similar EVs with diameters ranging from 30 to 200 nm, which contain complex populations of transcripts. RNA-seq identified abundant ribosomal RNAs, related pseudogenes and retrotransposons in human and Drosophila EVs. Vault RNAs and Y RNAs abounded in human samples, whereas small nucleolar RNAs involved in pseudouridylation were most prevalent in Drosophila EVs. Numerous mRNAs were identified, largely consisting of exonic sequences displaying full-length read coverage and enriched for translation and electronic transport chain functions. By analogy with human systems, these sizeable similarities suggest that EVs could potentially enable RNA-mediated intercellular communication in Drosophila.