Analysis of the structure and activity of the promoter regions of the metallothionein genes of the freshwater pearl mussel Hyriopsis schlegelii.

To investigate the regulation of metallothionein genes (HsMTs) of Hyriopsis schlegelii, 1,121-bp and 1,270-bp regions of the HsMT1 and HsMT2 promoters were cloned and analyzed, respectively. The two promoters shared partially conserved features and possessed distinct characteristics such as the number or position of metal response elements (MREs). Further analysis of the HsMT1 and HsMT2 promoters was performed by the reporter assay using the luciferase gene. Both promoters were activated by various metals, and presented different levels of metal ions inducibility in human hepatoblastoma cells. Deletion mutant assays demonstrated that both the longest promoter regions achieved the maximum inducibility, and the metal inducibility was dependent on the presence of the MRE in HsMT1 and the distal MRE in HsMT2. In addition, we cloned a putative metal responsive transcription factor (hereby designated as HsMTF-like) and studied its effect on HsMTs expression in human hepatoblastoma cells. An in vivo assay demonstrated that HsMTF-like activates basal HsMTs transcription level, and the MRE in the HsMTs promoter mediates this activation process. Moreover, this basal transcription level can be further boosted by zinc treatment. In conclusion, the regulation mechanism for MT activation in H. schlegelii should be evolutionarily conserved.

Investigation of Macroscopic Mechanical Behavior of Magnetorheological Elastomers under Shear Deformation Using Microscale Representative Volume Element Approach.

Magnetorheological elastomers (MREs) are a class of smart materials with rubber-like qualities, demonstrating revertible magnetic field-dependent viscoelastic properties, which makes them an ideal candidate for development of the next generation of adaptive vibration absorbers. This research study aims at the development of a finite element model using microscale representative volume element (RVE) approach to predict the field-dependent shear behavior of MREs. MREs with different elastomeric matrices, including silicone rubber Ecoflex 30 and Ecoflex 50, and carbonyl iron particles (CIPs) have been considered as magnetic particles. The stress-strain characteristic of the pure silicon rubbers was evaluated experimentally to formulate the nonlinear Ogden strain energy function to describe hyper-elastic behavior of the rubbery matrix. The obtained mechanical and magnetic properties of the matrix and inclusions were integrated into COMSOL Multiphysics to develop the RVE for the MREs, in 2D and 3D configurations, with CIP volume fraction varying from 5% to 40%. Periodic boundary condition (PBC) was imposed on the RVE boundaries, while undergoing shear deformation subjected to magnetic flux densities of 0-0.4 T. Comparing the results from 2D and 3D modeling of isotropic MRE-RVE with the experimental results from the literature suggests that the 3D MRE-RVE can be effectively used to accurately predict the influence of varying factors including matrix type, volume fraction of magnetic particles, and applied magnetic field on the mechanical behavior of MREs.

Analysis of an Adaptive Periodic Low-Frequency Wave Filter Featuring Magnetorheological Elastomers.

This study aims to enhance and tune wave-propagation properties (Bandgaps) of periodic structures featuring magnetorheological elastomers (MREs). For this purpose, first, a basic model of periodic structures (square unit cell with cross-shaped arms), which does not possess noise filtering properties in the conventional configuration, is considered. A passive attenuation zone is then proposed by adding a cylindrical core mass to the center of the conventional geometry and changing arm angles, which permitted new bandgap areas. It was shown that better wave-filtering performance may be achieved by introducing a large radius of the cylindrical core as well as low negative cross-arm angles. The modified configuration of the unit cell was subsequently utilized as the basic model for the development of magnetoactive metamaterial using a MRE capable of varying the bandgaps areas upon application of an external magnetic field. The finite element model of the proposed MRE-based periodic unit cell was developed, and the Bloch theorem was employed to systematically investigate the ability of the proposed adaptive periotic structure to attenuate low-frequency noise and vibration. Results show that the proposed MRE-based periodic wave filter can provide wide bandgap areas which can be adaptively changed and tuned using the applied magnetic field. The findings in this study can provide an essential guide for the development of novel adaptive periodic structures to filter low-frequency noises in the wide frequency band.

The prognostic miR-532-5p-correlated ceRNA-mediated lipid droplet accumulation drives nodal metastasis of cervical cancer.

Introduction: The prognosis for cervical cancer (CC) patients with lymph node metastasis (LNM) is extremely poor. Lipid droplets (LDs) have a pivotal role in promoting tumor metastasis. The crosstalk mechanism between LDs and LNM modulated in CC remains largely unknown. Objectives: This study aimed to construct a miRNA-dependent progonostic model for CC patients and investigate whether miR-532-5p has a biological impact on LNM by regualting LDs accumulation. Methods: LASSO-Cox regression was applied to establish a prognostic prediction model. miR-532-5p had the lowest P-value in RNA expression (P < 0.001) and prognostic prediction (P < 0.0001) and was selected for further study. The functional role of the prognostic miR-532-5p-correlated competing endogenous RNA (ceRNA) network was investigated to clarify the crosstalk between LDs and LNM. The underlying mechanism was determined using site-directed mutagenesis, dual luciferase reporter assays, RNA immunoprecipitation assays, and rescue experiments. A xenograft LNM model was established to evaluate the effect of miR-532-5p and orlistat combination therapy on tumor growth and LNM. Results: A novel 5-miRNAs prognostic signature was constructed to better predict the prognosis of CC patient. Further study demonstrated that miR-532-5p inhibited epithelial-mesenchymal transition and lymphangiogenesis by regulating LDs accumulation. Interestingly, we also found that LDs accumulation promoted cell metastasis in vitro. Mechanistically, we demonstrated a miR-532-5p-correlated ceRNA network in which LINC01410 was bound directly to miR-532-5p and effectively functioned as miR-532-5p sponge to disinhibit its target gene-fatty acid synthase (FASN). Combined therapy with miR-532-5p and FASN inhibitor-orlistat further inhibited tumor growth and LNM in vivo. Conclusion: Our findings highlight a LD accumulation-dependent mechanism of miR-532-5p-modulated LNM and support treatment with miR-532-5p/orlistat as novel strategy for treating patients with LNM in CC.

A cohort study of mild encephalitis/encephalopathy with a reversible splenial lesion in children.

PURPOSE: To investigate the clinical features, imaging features, and prognosis of mild encephalitis/encephalopathy with a reversible splenial lesion (MERS) in children METHODS: The clinical and imaging data of a cohort of 28 children diagnosed as MERS from January 2019 to October 2020 were retrospectively analyzed RESULTS: Of the 28 patients, 17 were males and 11 were females. The onset age ranged from 8 months to 12 years old, with an average age of 4 years and 2 months. All children developed normally before onset, and three of them had a history of febrile convulsion. More than half of the patients (62.9%) had preceding infections of gastrointestinal tract. All the cases developed seizures, and most (71.4%) had more than one time. Other neurological symptoms included dizziness/headache, consciousness disorder, limb weakness, blurred vision, and dysarthria. Cranial magnetic resonance imaging (MRI) showed lesions in the splenium of the corpus callosum in all, extending to other areas of the corpus callosum, bilateral semi-ovoid center, and adjacent periventricular in two cases. The clinical symptoms were relieved after steroids, intravenous immunogloblin, and symptomatic treatment, without abnormal neurodevelopment during the followed-up (2 months-2 years). Complete resolution of the lesions was observed 8-60 days after the initial MRI examinations CONCLUSION: MERS in children is related to prodromal infection mostly, with a wide spectrum of neurologic symptoms, characteristic MRI manifestations, and good prognosis.

Multiple Alu Exonization in 3'UTR of a Primate-Specific Isoform of CYP20A1 Creates a Potential miRNA Sponge.

Alu repeats contribute to phylogenetic novelties in conserved regulatory networks in primates. Our study highlights how exonized Alus could nucleate large-scale mRNA-miRNA interactions. Using a functional genomics approach, we characterize a transcript isoform of an orphan gene, CYP20A1 (CYP20A1_Alu-LT) that has exonization of 23 Alus in its 3'UTR. CYP20A1_Alu-LT, confirmed by 3'RACE, is an outlier in length (9 kb 3'UTR) and widely expressed. Using publically available data sets, we demonstrate its expression in higher primates and presence in single nucleus RNA-seq of 15,928 human cortical neurons. miRanda predicts ∼4,700 miRNA recognition elements (MREs) for ∼1,000 miRNAs, primarily originated within these 3'UTR-Alus. CYP20A1_Alu-LT could be a potential multi-miRNA sponge as it harbors ≥10 MREs for 140 miRNAs and has cytosolic localization. We further tested whether expression of CYP20A1_Alu-LT correlates with mRNAs harboring similar MRE targets. RNA-seq with conjoint miRNA-seq analysis was done in primary human neurons where we observed CYP20A1_Alu-LT to be downregulated during heat shock response and upregulated in HIV1-Tat treatment. In total, 380 genes were positively correlated with its expression (significantly downregulated in heat shock and upregulated in Tat) and they harbored MREs for nine expressed miRNAs which were also enriched in CYP20A1_Alu-LT. MREs were significantly enriched in these 380 genes compared with random sets of differentially expressed genes (P = 8.134e-12). Gene ontology suggested involvement of these genes in neuronal development and hemostasis pathways thus proposing a novel component of Alu-miRNA-mediated transcriptional modulation that could govern specific physiological outcomes in higher primates.

Characteristics of the competition among RNAs for the binding of shared miRNAs.

Competing endogenous RNAs (ceRNAs) are RNAs that share common miRNA binding sites and compete with each other for the miRNA association at these sites. The observation of this phenomenon in the cells altered the view of the miRNA target RNAs from molecules that are passively controlled by miRNAs to molecules that also modulate the miRNAs activity. In this review, we build a general profile of ceRNAS characteristics in order to facilitate ceRNAs identification by researchers. The information summarized here contains an actualized list of previously reported ceRNAs and classes of RNAs that can participate in this type of interaction, the expression behavior and characteristics of ceRNAs and miRNAs in the context of competition, the influence of the shared MREs/miRNAs numbers and the miRNA binding strength on the competition, reports on competition between RNAs in different subcellular localizations and the concept that ceRNAs may form a huge regulatory network in the cell.

Investigating miRNA-lncRNA Interactions: Computational Tools and Resources.

In the last two decades noncoding RNAs have been the recipients of increasing scientific interest. In particular, miRNAs, short (~22 nts) noncoding transcripts, have been thoroughly investigated since their essential role in posttranscriptional gene expression regulation had been established in the early 2000s. With the advent and the advancements of high-throughput sequencing technologies in recent years, long noncoding RNAs have also started to emerge as important actors in cellular functions and processes. Such transcripts, on average longer than 200 nt, whose functions have yet to be fully characterized, have recently been identified as regulatory elements of the RNAi pathway, harboring several miRNA response elements, uncovering the phenomena of competing endogenous RNAs (ceRNAs), or "sponge RNAs." The present chapter aims to provide a brief update on the actual biomedical relevance of ceRNAs, together with a summary of resources, tools, and practical examples of their application to aid researchers in the discovery and further elucidation of lncRNA-miRNA interactions.

A Bioinformatics Method for the Design of Live Attenuated Virus Vaccine Utilizing Host MicroRNA Response Elements.

The host microRNA machinery has been employed to control viral replication. To improve safety for live attenuated virus vaccines, the binding sites of the host microRNAs, so-called microRNA response elements (MREs), were incorporated into the virus sequences. These MREs were typically designed for a specific host microRNA and virus sequence with the effectiveness evaluated by experimental trials. Here, we describe a computational flow that can be used to simultaneously design and prioritize the effective MREs in large-scale.

SMYD1 and G6PD modulation are critical events for miR-206-mediated differentiation of rhabdomyosarcoma.

Rhadomyosarcoma (RMS) is the most common soft tissue sarcoma of childhood. RMS cells resemble fetal myoblasts but are unable to complete myogenic differentiation. In previous work we showed that miR-206, which is low in RMS, when induced in RMS cells promotes the resumption of differentiation by modulating more than 700 genes. To better define the pathways involved in the conversion of RMS cells into their differentiated counterpart, we focused on 2 miR-206 effectors emerged from the microarray analysis, SMYD1 and G6PD. SMYD1, one of the most highly upregulated genes, is a H3K4 histone methyltransferase. Here we show that SMYD1 silencing does not interfere with the proliferative block or with the loss anchorage independence imposed by miR-206, but severely impairs differentiation of ERMS, ARMS, and myogenic cells. Thus SMYD1 is essential for the activation of muscle genes. Conversely, among the downregulated genes, we found G6PD, the enzyme catalyzing the rate-limiting step of the pentose phosphate shunt. In this work, we confirmed that G6PD is a direct target of miR-206. Moreover, we showed that G6PD silencing in ERMS cells impairs proliferation and soft agar growth. However, G6PD overexpression does not interfere with the pro-differentiating effect of miR-206, suggesting that G6PD downmodulation contributes to - but is not an absolute requirement for - the tumor suppressive potential of miR-206. Targeting cancer metabolism may enhance differentiation. However, therapeutic inhibition of G6PD is encumbered by side effects. As an alternative, we used DCA in combination with miR-206 to increase the flux of pyruvate into the mitochondrion by reactivating PDH. DCA enhanced the inhibition of RMS cell growth induced by miR-206, and sustained it upon miR-206 de-induction. Altogether these results link miR-206 to epigenetic and metabolic reprogramming, and suggest that it may be worth combining differentiation-inducing with metabolism-directed approaches.

Competing endogenous RNAs (ceRNAs): new entrants to the intricacies of gene regulation.

The discovery of microRNAs (miRNAs) has led to a paradigm shift in our basic understanding of gene regulation. Competing endogenous RNAs (ceRNAs) are the recent entrants adding to the complexities of miRNA mediated gene regulation. ceRNAs are RNAs that share miRNA recognition elements (MREs) thereby regulating each other. It is apparent that miRNAs act as rheostats that fine-tune gene expression and maintain the functional balance of various gene networks. Thus MREs in coding and non-coding transcripts have evolved to become the crosstalk hubs of gene interactions, affecting the expression levels and activities of different ceRNAs. Decoding the crosstalk between MREs mediated by ceRNAs is critical to delineate the intricacies in gene regulation, and we have just begun to unravel this complexity.

Long non-coding RNAs: a new frontier in the study of human diseases.

With the development of whole genome and transcriptome sequencing technologies, long noncoding RNAs (lncRNAs) have received increased attention. Multiple studies indicate that lncRNAs act not only as the intermediary between DNA and protein but also as important protagonists of cellular functions. LncRNAs can regulate gene expression in many ways, including chromosome remodeling, transcription and post-transcriptional processing. Moreover, the dysregulation of lncRNAs has increasingly been linked to many human diseases, especially in cancers. Here, we reviewed the rapidly advancing field of lncRNAs and described the relationship between the dysregulation of lncRNAs and human diseases, highlighting the specific roles of lncRNAs in human diseases.