BackgroundRecently, thousands of circular RNAs (circRNAs) have been discovered in various tissues and cell types from human, mouse, fruit fly and nematodes. However, expression of circRNAs across mammalian brain development has never been examined.ResultsHere we profile the expression of circRNA in five brain tissues at up to six time-points during fetal porcine development, constituting the first report of circRNA in the brain development of a large animal. An unbiased analysis reveals a highly complex regulation pattern of thousands of circular RNAs, with a distinct spatio-temporal expression profile. The amount and complexity of circRNA expression was most pronounced in cortex at day 60 of gestation. At this time-point we find 4634 unique circRNAs expressed from 2195 genes out of a total of 13,854 expressed genes. Approximately 20 % of the porcine splice sites involved in circRNA production are functionally conserved between mouse and human. Furthermore, we observe that “hot-spot” genes produce multiple circRNA isoforms, which are often differentially expressed across porcine brain development. A global comparison of porcine circRNAs reveals that introns flanking circularized exons are longer than average and more frequently contain proximal complementary SINEs, which potentially can facilitate base pairing between the flanking introns. Finally, we report the first use of RNase R treatment in combination with in situ hybridization to show dynamic subcellular localization of circRNA during development.ConclusionsThese data demonstrate that circRNAs are highly abundant and dynamically expressed in a spatio-temporal manner in porcine fetal brain, suggesting important functions during mammalian brain development.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-015-0801-3) contains supplementary material, which is available to authorized users.

The hSuv3 (human Suv3) helicase has been shown to be a major player in mitochondrial RNA surveillance and decay, but its physiological role might go beyond this functional niche. hSuv3 has been found to interact with BLM (Bloom’s syndrome protein) and WRN (Werner’s syndrome protein), members of the RecQ helicase family involved in multiple DNA metabolic processes, and in protection and stabilization of the genome. In the present study, we have addressed the possible role of hSuv3 in genome maintenance by examining its potential association with key interaction partners of the RecQ helicases. By analysis of hSuv3 co-IP (co-immunoprecipitation) complexes, we identify two new interaction partners of hSuv3: the RPA (replication protein A) and FEN1 (flap endonuclease 1). Utilizing an in vitro biochemical assay we find that low amounts of RPA inhibit helicase activity of hSuv3 on a forked substrate. Another single-strand-binding protein, mtSSB (mitochondrial single-strand-binding protein), fails to affect hSuv3 activity, indicating that the functional interaction is specific for hSuv3 and RPA. Further in vitro studies demonstrate that the flap endonuclease activity of FEN1 is stimulated by hSuv3 independently of flap length. hSuv3 is generally thought to be a mitochondrial helicase, but the physical and functional interactions between hSuv3 and known RecQ helicase-associated proteins strengthen the hypothesis that hSuv3 may play a significant role in nuclear DNA metabolism as well.

The porcine brain closely resembles the human brain in aspects such as development and morphology. Temporal miRNA profiling in the developing embryonic porcine cortex revealed a distinct set of miRNAs, including miR-34c and miR-204, which exhibited a highly specific expression profile across the time of cortical folding. These miRNAs were found to target Doublecortin (DCX), known to be involved in neuron migration during cortical folding of gyrencephalic brains. In vivo modulation of miRNA expression in mouse embryos confirmed that miR-34c and miR-204 can control neuronal migration and cortical morphogenesis, presumably by posttranscriptional regulation of DCX.

Receptor- and adsorptive-mediated transport through brain endothelial cells (BEC) of the blood-brain barrier (BBB) involves a complex array of subcellular vesicular structures, the endo-lysosomal system. It consists of several types of vesicles, such as early, recycling, and late endosomes, retromer-positive structures, and lysosomes. Since this system is important for receptor-mediated transcytosis of drugs across brain capillaries, our aim was to characterise the endo-lysosomal system in BEC with emphasis on their interactions with astrocytes. We used primary porcine BEC in monoculture and in co-culture with primary rat astrocytes. The presence of astrocytes changed the intraendothelial vesicular network and significantly impacted vesicular number, morphology, and distribution. Additionally, gene set enrichment analysis revealed that 60 genes associated with vesicular trafficking showed altered expression in co-cultured BEC. Cytosolic proteins involved in subcellular trafficking were investigated to mark transport routes, such as RAB25 for transcytosis. Strikingly, the adaptor protein called AP1-μ1B, important for basolateral sorting in epithelial cells, was not expressed in BEC. Altogether, our data pin-point unique features of BEC trafficking network, essentially mapping the endo-lysosomal system of in vitro BBB models. Consequently, our findings constitute a valuable basis for planning the optimal route across the BBB when advancing drug delivery to the brain.