In Human Pseudouridine Synthase 1 (hPus1), a C-Terminal Helical Insert Blocks tRNA from Binding in the Same Orientation as in the Pus1 Bacterial Homologue TruA,Consistent with Their Different Target Selectivities

Human pseudouridine (Ψ) synthase Pus1 (hPus1) modifies specific uridine residues in several non-coding RNAs: tRNA, U2 spliceosomal RNA, and steroid receptor activator RNA. We report three structures of the catalytic core domain of hPus1 from two crystal forms, at 1.8 Å resolution. The structures are the first of a mammalian Ψ synthase from the set of five Ψ synthase families common to all kingdoms of life. hPus1 adopts a fold similar to bacterial Ψ synthases, with a central antiparallel β-sheet flanked by helices and loops. A flexible hinge at the base of the sheet allows the enzyme to open and close around an electropositive active-site cleft. In one crystal form, a molecule of Mes [2-(N-morpholino)ethane sulfonic acid] mimics the target uridine of an RNA substrate. A positively charged electrostatic surface extends from the active site towards the N-terminus of the catalytic domain, suggesting an extensive binding site specific for target RNAs. Two α-helices C-terminal to the core domain, but unique to hPus1, extend along the back and top of the central β-sheet and form the walls of the RNA binding surface. Docking of tRNA to hPus1 in a productive orientation requires only minor conformational changes to enzyme and tRNA. The docked tRNA is bound by the electropositive surface of the protein employing a completely different binding mode than that seen for the tRNA complex of the Escherichia coli homologue TruA.     

Detection of long non-coding RNAs in human breastmilk extracellular vesicles: Implications for early child development

Breastmilk has many documented beneficial effects on the developing human infant, but the components of breastmilk that influence these developmental pathways have not been fully elucidated. Increasing evidence suggests that non-coding RNAs encapsulated in extracellular vesicles (EVs) represent an important mechanism of communication between the mother and child. Long non-coding RNAs (lncRNAs) are of particular interest given their key role in gene expression and development. However, it is not known whether breastmilk EVs contain lncRNAs. We used qRT-PCR to determine whether EVs isolated from human breastmilk contain lncRNAs previously reported to be important for developmental processes. We detected 55 of the 87 screened lncRNAs in EVs from the 30 analyzed breastmilk samples, and CRNDE, DANCR, GAS5, SRA1 and ZFAS1 were detected in >90% of the samples. GAS5, SNHG8 and ZFAS1 levels were highly correlated (Spearman's rho > 0.9; P < 0.0001), which may indicate that the loading of these lncRNAs into breastmilk EVs is regulated by the same pathways. The detected lncRNAs are important epigenetic regulators involved in processes such as immune cell regulation and metabolism. They may target a repertoire of recipient cells in offspring and could be essential for child development and health. Further experimental and epidemiological studies are warranted to determine the impact of breastmilk EV-encapsulated lnRNAs in mother to child signaling.     

RNA‐Seq bulked segregant analysis enables the identification of high‐resolution genetic markers for breeding in hexaploid wheat

The identification of genetic markers linked to genes of agronomic importance is a major aim of crop research and breeding programmes. Here, we identify markers for Yr15, a major disease resistance gene for wheat yellow rust, using a segregating F₂ population. After phenotyping, we implemented RNA sequencing (RNA‐Seq) of bulked pools to identify single‐nucleotide polymorphisms (SNP) associated with Yr15. Over 27 000 genes with SNPs were identified between the parents, and then classified based on the results from the sequenced bulks. We calculated the bulk frequency ratio (BFR) of SNPs between resistant and susceptible bulks, selecting those showing sixfold enrichment/depletion in the corresponding bulks (BFR > 6). Using additional filtering criteria, we reduced the number of genes with a putative SNP to 175. The 35 SNPs with the highest BFR values were converted into genome‐specific KASP assays using an automated bioinformatics pipeline (PolyMarker) which circumvents the limitations associated with the polyploid wheat genome. Twenty‐eight assays were polymorphic of which 22 (63%) mapped in the same linkage group as Yr15. Using these markers, we mapped Yr15 to a 0.77‐cM interval. The three most closely linked SNPs were tested across varieties and breeding lines representing UK elite germplasm. Two flanking markers were diagnostic in over 99% of lines tested, thus providing a reliable haplotype for marker‐assisted selection in these breeding programmes. Our results demonstrate that the proposed methodology can be applied in polyploid F₂ populations to generate high‐resolution genetic maps across target intervals.     

MBD结构域和SRA结构域识别甲基化DNA的结构机理

DNA胞嘧啶5-甲基化修饰是表观遗传重要的修饰之一,其对基因表达的调控依赖下游的识别蛋白识别和传递甲基化信号.本文围绕两种主要的甲基化DNA识别结构域——MBD结构域和SRA结构域,综述了它们识别不同修饰形式DNA的结构基础以及发挥功能的分子机理.