Pre-announcement of symbiotic guests: transcriptional reprogramming by mycorrhizal lipochitooligosaccharides shows a strict co-dependency on the GRAS transcription factors NSP1 and RAM1

BACKGROUND: More than 80 % of all terrestrial plant species establish an arbuscular mycorrhiza (AM) symbiosis with Glomeromycota fungi. This plant-microbe interaction primarily improves phosphate uptake, but also supports nitrogen, mineral, and water aquisition. During the pre-contact stage, the AM symbiosis is controled by an exchange of diffusible factors from either partner. Amongst others, fungal signals were identified as a mix of sulfated and non-sulfated lipochitooligosaccharides (LCOs), being structurally related to rhizobial nodulation (Nod)-factor LCOs that in legumes induce the formation of nitrogen-fixing root nodules. LCO signals are transduced via a common symbiotic signaling pathway (CSSP) that activates a group of GRAS transcription factors (TFs). Using complex gene expression fingerprints as molecular phenotypes, this study primarily intended to shed light on the importance of the GRAS TFs NSP1 and RAM1 for LCO-activated gene expression during pre-symbiotic signaling. RESULTS: We investigated the genome-wide transcriptional responses in 5 days old primary roots of the Medicago truncatula wild type and four symbiotic mutants to a 6 h challenge with LCO signals supplied at 10(-7/-8) M. We were able to show that during the pre-symbiotic stage, sulfated Myc-, non-sulfated Myc-, and Nod-LCO-activated gene expression almost exclusively depends on the LysM receptor kinase NFP and is largely controled by the CSSP, although responses independent of this pathway exist. Our results show that downstream of the CSSP, gene expression activation by Myc-LCOs supplied at 10(-7/-8) M strictly required both the GRAS transcription factors RAM1 and NSP1, whereas those genes either co- or specifically activated by Nod-LCOs displayed a preferential NSP1-dependency. RAM1, a central regulator of root colonization by AM fungi, controled genes activated by non-sulfated Myc-LCOs during the pre-symbiotic stage that are also up-regulated in areas with early physical contact, e.g. hyphopodia and infecting hyphae; linking responses to externally applied LCOs with early root colonization. CONCLUSIONS: Since both RAM1 and NSP1 were essential for the pre-symbiotic transcriptional reprogramming by Myc-LCOs, we propose that downstream of the CSSP, these GRAS transcription factors act synergistically in the transduction of those diffusible signals that pre-announce the presence of symbiotic fungi.DFG/SPP1212/KU-1478/4-1DFG/SPP1212/KU-1478/4-4DFG/SPP1212/KU-1478/4-2DFG/SPP1212/KU-1478/4-

Profiling and Validation of the Circular RNA Repertoire in Adult Murine Hearts

For several decades, cardiovascular disease has been the leading cause of death throughout all countries. There is a strong genetic component to many disease subtypes (e.g., cardiomyopathy) and we are just beginning to understand the relevant genetic factors. Several studies have related RNA splicing to cardiovascular disease and circular RNAs (circRNAs) are an emerging player. circRNAs, which originate through back-splicing events from primary transcripts, are resistant to exonucleases and typically not polyadenylated. Initial functional studies show clear phenotypic outcomes for selected circRNAs. We provide, for the first time, a comprehensive catalogue of RNase R-resistant circRNA species for the adult murine heart. This work combines state-of-the-art circle sequencing with our novel DCC software to explore the circRNA landscape of heart tissue. Overall, we identified 575 circRNA species that pass a beta-binomial test for enrichment (false discovery rate of 1%) in the exonuclease-treated sequencing sample. Several circRNAs can be directly attributed to host genes that have been previously described as associated with cardiovascular disease. Further studies of these candidate circRNAs may reveal disease-relevant properties or functions of specific circRNAs

Corresponding authors Distributed under Creative Commons CC-BY 4.0 FUCHS-towards full circular RNA characterization using RNAseq

ABSTRACT Circular RNAs (circRNAs) belong to a recently re-discovered species of RNA that emerge during RNA maturation through a process called back-splicing. A downstream 5 splice site is linked to an upstream 3 splice site to form a circular transcript instead of a canonical linear transcript. Recent advances in next-generation sequencing (NGS) have brought circRNAs back into the focus of many scientists. Since then, several studies reported that circRNAs are differentially expressed across tissue types and developmental stages, implying that they are actively regulated and not merely a byproduct of splicing. Though functional studies have shown that some circRNAs could act as miRNA-sponges, the function of most circRNAs remains unknown. To expand our understanding of possible roles of circular RNAs, we propose a new pipeline that could fully characterizes candidate circRNA structure from RNAseq data-FUCHS: FUll CHaracterization of circular RNA using RNA-Sequencing. Currently, most computational prediction pipelines use back-spliced reads to identify circular RNAs. FUCHS extends this concept by considering all RNA-seq information from long reads (typically >150 bp) to learn more about the exon coverage, the number of double break point fragments, the different circular isoforms arising from one host-gene, and the alternatively spliced exons within the same circRNA boundaries. This new knowledge will enable the user to carry out differential motif enrichment and miRNA seed analysis to determine potential regulators during circRNA biogenesis. FUCHS is an easy-to-use Python based pipeline that contributes a new aspect to the circRNA research