Project description:microRNAs are frequently modified by addition of untemplated nucleotides to the 3' end, but the role of this tailing is often unclear. Here we characterize the prevalence and functional consequences of microRNA tailing in vivo, using Caenorhabditis elegans. MicroRNA tailing in C. elegans consists mostly of mono-uridylation of mature microRNA species, with rarer mono-adenylation which is likely added to microRNA precursors. Through a targeted RNAi screen, we discover that the TUT4/TUT7 gene family member CID-1/CDE-1/PUP-1 is required for uridylation, whereas the GLD2 gene family member F31C3.2-here named GLD-2-related 2 (GLDR-2)-is required for adenylation. Thus, the TUT4/TUT7 and GLD2 gene families have broadly conserved roles in miRNA modification. We specifically examine the role of tailing in microRNA turnover. We determine half-lives of microRNAs after acute inactivation of microRNA biogenesis, revealing that half-lives are generally long (median = 20.7 h), as observed in other systems. Although we observe that the proportion of tailed species increases over time after biogenesis, disrupting tailing does not alter microRNA decay. Thus, tailing is not a global regulator of decay in C. elegans. Nonetheless, by identifying the responsible enzymes, this study lays the groundwork to explore whether tailing plays more specialized context- or miRNA-specific regulatory roles.
Project description:Many microRNAs (miRNAs) exist alongside abundant miRNA isoforms (isomiRs), most of which arise from post-maturation sequence modifications, such as 3’ uridylation and adenylation. However, the ways in which these sequence modifications affect miRNA function remain poorly understood. To this aim, we have generated single knock-out cell lines of TUT4, TUT7 and TENT2 (TUT2), as well as double knock-out (DKO) and triple knock-out (TKO) cell lines. Here, using these different cell lines, we have discovered that some of the redundant functions and specific functions of each tailing enzyme. Our study provides a comprehensive characterization of tailing on miRNAs.
Project description:To analyse the impact of URT1-mediated uridylation on miRNA and siRNA tailing, we deep-sequenced small RNA libraries for WT and urt1 duplicate samples at the same developmental stage that was analyzed by TAIL-seq, i.e., two-week-old seedlings. Examination of miRNA and siRNA tailing in WT and urt1 samples.
Project description:Potash tailing piles located in Germany represent extremely hypersaline locations that negatively affect neighbouring environments and limit the development of higher vegetation. However, biocrusts, as cryptogamic covers, inhabit some of these areas and provide essential ecological functions, but, nevertheless, they remain poorly described. Here, we applied high-throughput sequencing (HTS) and targeted four groups of microorganisms: bacteria, cyanobacteria, fungi and other eukaryotes. The sequencing of the 16S rRNA gene revealed the dominance of Proteobacteria, Cyanobacteria and Actinobacteria. Additionally, we applied yanobacteria-specific primers for a detailed assessment of the cyanobacterial community, which was dominated by members of the filamentous orders Synechococcales and Oscillatoriales. Furthermore, the majority of reads in the studied biocrusts obtained by sequencing of the 18S rRNA gene belonged to eukaryotic microalgae. In addition, sequencing of the internal rDNA transcribed spacer region (ITS) showed the dominance of Ascomycota within the fungal community. Overall, these molecular data provided the first detailed overview of microorganisms associated with biocrusts inhabiting highly saline potash tailing piles and showed the dissimilarities in microbial diversity among the samples.