Expansion of MIR482/2118 by a class‐II transposable element in cotton

Some plant microRNA (miRNA) families contain multiple members generating identical or highly similar mature miRNA variants. Mechanisms underlying the expansion of miRNA families remain elusive, although tandem and/or segmental duplications have been proposed. In this study of two tetraploid cottons, Gossypium hirsutum and Gossypium barbadense, and their extant diploid progenitors, Gossypium arboreum and Gossypium raimondii, we investigated the gain and loss of members of the miR482/2118 superfamily, which modulates the expression of nucleotide‐binding site leucine‐rich repeat (NBS‐LRR) disease resistance genes. We found significant expansion of MIR482/2118d in G. barbadense, G. hirsutum and G. raimondii, but not in G. arboreum. Several newly expanded MIR482/2118d loci have mutated to produce different miR482/2118 variants with altered target‐gene specificity. Based on detailed analysis of sequences flanking these MIR482/2118 loci, we found that this expansion of MIR482/2118d and its derivatives resulted from an initial capture of an MIR482/2118d by a class‐II DNA transposable element (TE) in G. raimondii prior to the tetraploidization event, followed by transposition to new genomic locations in G. barbadense, G. hirsutum and G. raimondii. The ‘GosTE’ involved in the capture and proliferation of MIR482/2118d and its derivatives belongs to the PIF/Harbinger superfamily, generating a 3‐bp target site duplication upon insertion at new locations. All orthologous MIR482/2118 loci in the two diploids were retained in the two tetraploids, but mutation(s) in miR482/2118 were observed across all four species as well as in different cultivars of both G. barbadense and G. hirsutum, suggesting a dynamic co‐evolution of miR482/2118 and its NBS‐LRR targets. Our results provide fresh insights into the mechanisms contributing to MIRNA proliferation and enrich our knowledge on TEs.     

Ca2+-dependent mechanism of membrane insertion and destabilization by the SARS-CoV-2 fusion peptide

Cell penetration after recognition of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus by the ACE2 receptor and the fusion of its viral envelope membrane with cellular membranes are the early steps of infectivity. A region of the Spike protein of the virus, identified as the “fusion peptide” (FP), is liberated at its N-terminal site by a specific cleavage occurring in concert with the interaction of the receptor-binding domain of the Spike. Studies have shown that penetration is enhanced by the required binding of Ca²⁺ ions to the FPs of coronaviruses, but the mechanisms of membrane insertion and destabilization remain unclear. We have predicted the preferred positions of Ca²⁺ binding to the SARS-CoV-2-FP, the role of Ca²⁺ ions in mediating peptide-membrane interactions, the preferred mode of insertion of the Ca²⁺-bound SARS-CoV-2-FP, and consequent effects on the lipid bilayer from extensive atomistic molecular dynamics simulations and trajectory analyses. In a systematic sampling of the interactions of the Ca²⁺-bound peptide models with lipid membranes, SARS-CoV-2-FP penetrated the bilayer and disrupted its organization only in two modes involving different structural domains. In one, the hydrophobic residues F833/I834 from the middle region of the peptide are inserted. In the other, more prevalent mode, the penetration involves residues L822/F823 from the LLF motif, which is conserved in CoV-2-like viruses, and is achieved by the binding of Ca²⁺ ions to the D830/D839 and E819/D820 residue pairs. FP penetration is shown to modify the molecular organization in specific areas of the bilayer, and the extent of membrane binding of the SARS-CoV-2 FP is significantly reduced in the absence of Ca²⁺ ions. These findings provide novel mechanistic insights regarding the role of Ca²⁺ in mediating SARS-CoV-2 fusion and provide a detailed structural platform to aid the ongoing efforts in rational design of compounds to inhibit SARS-CoV-2 cell entry.     

A multiscale model for multiple platelet aggregation in shear flow

Biomechanics and Modeling in Mechanobiology - We developed a multiscale model for simulating aggregation of multiple, free-flowing platelets in low–intermediate shear viscous flow, in which...     

Large Scale Isolation of Pig Muscle Phosphoglucose Isomerase

large-scale purification procedure for phosphoglucose isomerase from pig skeletal muscle is described. It consists of two fractionations by selective precipitation and two ion exchange chromatography steps yielding an end product of approximately 900 units (micromoles of sub-strate converted to product per rain per mg of protein, at 30°) specific activity. The method separates three isoenzymic forms with an overall recovery of about 30% of the original total enzyme activity in the form of Isoenzyme III, the latter being the predominant enzyme species.     

REACTION OF ANHYDRONUCLEOSIDES WITH MAGNESIUM ALKOXIDES: REGIOSPECIFIC SYNTHESIS OF 2′-O-ALKYLPYRIMIDINE NUCLEOSIDES.

Abstract

A novel approach to 2′-O-alkylpyrimidine nucleosides involving a 3′- hydroxyl assisted intramolecular delivery of a divalent metal alkoxide leads to a regiospecific opening of the anhydropyrimidine linkage at the 2′-position. Thus, reaction of 5′-protected 2,2′-anhydrouridine with magnesium or calcium alkoxides in DMF affords exclusively the corresponding 2′-O-alkyluridines in reasonable yields.     

Dendritic Branch Intersections Are Structurally Regulated Targets for Efficient Axonal Wiring and Synaptic Clustering

Synaptic clustering on dendritic branches enhances plasticity, input integration and neuronal firing. However, the mechanisms guiding axons to cluster synapses at appropriate sites along dendritic branches are poorly understood. We searched for such a mechanism by investigating the structural overlap between dendritic branches and axons in a simplified model of neuronal networks - the hippocampal cell culture. Using newly developed software, we converted images of meshes of overlapping axonal and dendrites into topological maps of intersections, enabling quantitative study of overlapping neuritic geometry at the resolution of single dendritic branch-to-branch and axon-to-branch crossings. Among dendro-dendritic crossing configurations, it was revealed that the orientations through which dendritic branches cross is a regulated attribute. While crossing angle distribution among branches thinner than 1 µm appeared to be random, dendritic branches 1 µm or wider showed a preference for crossing each other at angle ranges of either 50°–70° or 80°–90°. It was then found that the dendro-dendritic crossings themselves, as well as their selective angles, both affected the path of axonal growth. Axons displayed 4 fold stronger tendency to traverse within 2 µm of dendro-dendritic intersections than at farther distances, probably to minimize wiring length. Moreover, almost 70% of the 50°–70° dendro-denritic crossings were traversed by axons from the obtuse angle’s zone, whereas only 15% traversed through the acute angle’s zone. By contrast, axons showed no orientation restriction when traversing 80°–90° crossings. When such traverse behavior was repeated by many axons, they converged in the vicinity of dendro-dendritic intersections, thereby clustering their synaptic connections. Thus, the vicinity of dendritic branch-to-branch crossings appears to be a regulated structure used by axons as a target for efficient wiring and as a preferred site for synaptic clustering. This synaptic clustering mechanism may enhance synaptic co-activity and plasticity.     

Mutant p53 Attenuates the Anti-Tumorigenic Activity of Fibroblasts-Secreted Interferon Beta

Mutations in the p53 tumor suppressor protein are highly frequent in tumors and often endow cells with tumorigenic capacities. We sought to examine a possible role for mutant p53 in the cross-talk between cancer cells and their surrounding stroma, which is a crucial factor affecting tumor outcome. Here we present a novel model which enables individual monitoring of the response of cancer cells and stromal cells (fibroblasts) to co-culturing. We found that fibroblasts elicit the interferon beta (IFNβ) pathway when in contact with cancer cells, thereby inhibiting their migration. Mutant p53 in the tumor was able to alleviate this response via SOCS1 mediated inhibition of STAT1 phosphorylation. IFNβ on the other hand, reduced mutant p53 RNA levels by restricting its RNA stabilizer, WIG1. These data underscore mutant p53 oncogenic properties in the context of the tumor microenvironment and suggest that mutant p53 positive cancer patients might benefit from IFNβ treatment.