Tightly coordinated, reciprocal embryo-maternal interactions affect gene expression during early pregnancy. Recently, microRNAs (miRNAs) have emerged as new players in the fine tuning of embryo development and implantation in mammals via posttranscriptional gene regulation mechanisms. Here, we integrated transcriptomic and computational approaches to profile miRNAs and miRNA synthesis and transport-related genes at different developmental stages of porcine conceptuses and trophoblast during early pregnancy in the pig. Using semiquantitative RT-PCR, we examined mRNA levels of 10 genes encoding proteins involved in miRNA synthesis and transport: DROSHA, DGCR8, XPO5, DICER1, TARBP2, TNRC6A, AGO1, AGO2, AGO3, and AGO4. Custom, multispecies microarrays were used to profile miRNAs. Prediction algorithms of miRNA-mRNA interactions allowed identification of target transcripts for the analyzed miRNAs. These included VEGF, LIF, PTGS2, and IL-6R, known to be crucial components of embryo-maternal interactions in the pig. Two selected miRNAs, miR-26a and miR-125b, were tested for the presence in the extracellular vesicles isolated from uterine luminal flushings during pregnancy. Results of in vitro study demonstrated that miRNAs, such as miR-125b, can regulate expression of genes crucial for embryo development and implantation in porcine endometrial luminal epithelial cells. For the first time, expression profiles of miRNAs and related genes in porcine conceptuses and trophoblast during maternal recognition of pregnancy and embryo implantation in the pig were described. Altogether, our results indicate potential roles of these small, noncoding RNAs in the early development of embryos and embryo-maternal cross-talk during early pregnancy in the pig.

BackgroundEmbryo implantation is a complex, synchronized process that requires establishment of a reciprocal dialogue between a receptive endometrium and developing blastocysts. Recently, microRNAs (miRNAs), known to modulate gene expression through post-transcriptional mechanisms, were implicated in regulation of early pregnancy events including maternal recognition of pregnancy and implantation. To characterize complex transcriptomic changes, expression of miRNAs in pregnant and cyclic endometria collected on days 12, 16 and 20 was analyzed using Illumina deep sequencing and analyzed with bioinformatic pipeline. Moreover, expression profiles of ten genes related to miRNA synthesis and transport such as DROSHA, DGCR8, XPO5, DICER, TARBP2, TNRC6A, and AGO1-4 were determined.ResultsAmong genes involved in miRNA transport and synthesis DROSHA, XPO5, DICER1, TARBP, and AGO1 expression was affected by the reproductive status. Moreover, DICER1 and AGO2 proteins were localized in luminal and glandular epithelium with immunofluorescence staining. Several hundred mature, canonical and non-canonical miRNAs were found to be expressed in the endometrial samples. Detailed analysis revealed that miRNA length variants, isomiRs, accounted for the vast majority of defined sequences. Both miRNA and isomiR of miR-140-3p were shown to affect expression of putative targets in endometrial stromal cells in vitro. Computational analysis of putative target genes for miRNAs differentially expressed (DE) between pregnant and cyclic animals resulted in lists of biological processes and regulatory pathways indicating their role in cellular development, cell cycle, immunological response and organismal development. Among predicted target genes for DE miRNAs, vascular endothelial growth factor (VEGF), progesterone and estradiol receptors (PGR, ESR1) and leukemia inhibitory factor (LIF) were found.ConclusionsThis research revealed a repertoire of pregnancy-related miRNAs in porcine endometrium during initial stages of conceptus implantation and during the estrous cycle, and sheds light on mechanisms regulating miRNA-mediated gene expression at the maternal-conceptus interface.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2172-2) contains supplementary material, which is available to authorized users.

The assortment of cellular microRNAs (“microRNAome”) is a vital readout of cellular homeostasis, but the mechanisms that regulate the microRNAome are poorly understood. The microRNAome of glioblastoma is substantially down-regulated in comparison to the normal brain. Here, we find malfunction of the posttranscriptional maturation of the glioblastoma microRNAome and link it to aberrant nuclear localization of DICER, the major enzymatic complex responsible for microRNA maturation. Analysis of DICER’s nuclear interactome reveals the presence of an RNA binding protein, RBM3, and of a circular RNA, circ2082, within the complex. Targeting of this complex by knockdown of circ2082 results in the restoration of cytosolic localization of DICER and widespread derepression of the microRNAome, leading to transcriptome-wide rearrangements that mitigate the tumorigenicity of glioblastoma cells in vitro and in vivo with correlation to favorable outcomes in patients with glioblastoma. These findings uncover the mechanistic foundation of microRNAome deregulation in malignant cells.