Four-way Junction Research Articles

FebRNA: An automated fragment-ensemble-based model for building RNA 3D structures

Knowledge of RNA three-dimensional (3D) structures is critical to understanding the important biological functions of RNAs. Although various structure prediction models have been developed, the high-accuracy predictions of RNA 3D structures are still limited to the RNAs with short lengths or with simple topology. In this work, we proposed a new model, namely FebRNA, for building RNA 3D structures through fragment assembly based on coarse-grained (CG) fragment ensembles. Specifically, FebRNA is composed of four processes: establishing the library of different types of non-redundant CG fragment ensembles regardless of the sequences, building CG 3D structure ensemble through fragment assembly, identifying top-scored CG structures through a specific CG scoring function, and rebuilding the all-atom structures from the top-scored CG ones. Extensive examination against different types of RNA structures indicates that FebRNA consistently gives the reliable predictions on RNA 3D structures, including pseudoknots, three-way junctions, four-way and five-way junctions, and RNAs in the RNA-Puzzles. FebRNA is available on the Web site: https://github.com/Tan-group/FebRNA.

Nuclease resistance and protein recognition properties of DNA and hybrid PNA-DNA four-way junctions

Nucleic acids possess unique biochemical features that make them ideal candidates to inhibit “difficult to target” proteins. The limited stability of nucleic acids in vivo presents a major obstacle to their development as drugs. Here, immobile four-way junctions (4WJs) are used to target the DNA-binding cytokine, High Mobility Group B1. Hybrid 4WJs composed of DNA and peptide nucleic acids (PNA) are investigated. PNA possess enhanced nuclease stability vs. DNA. 4WJs are incubated with Exonuclease III and DNase I. The nuclease assays show that 4WJs containing multiple PNAs possess significantly higher stability. Circular dichroism assays are used to probe the groove topology of 4WJs with the minor groove binder, DAPI. The CD data indicates that multi-PNA 4WJs possess altered minor groove dimensions that reduces DAPI binding affinity. Logic suggests that the minor groove of multi-PNA hybrids possess significant perturbations to the topology and local electrostatic environment that prevents proper binding/recognition by nucleases and thus enhances stability.

Traffic Prediction for Intelligent Transportation Systems using Machine Learning

Over the past few decades, ITS have spiked an increasing research interest as a promising discipline for revolutionizing the transportation sector and solving common traffic and vehicle-related problems. ITS comprise a multitude of interconnected engineering feats that function as an entity for optimizing network-scale travel experiences from a technical, social, economic, and environmental aspect. Such optimizations necessitate the advancement of information and communication technologies, electronic sensors, control systems, and computers, which high-lights the data-driven nature of modern ITS. In this paper we design a system which uses machine learning algorithm using SVM, KNN and CNN algorithm which is a novel system which will provide intelligence to the current traffic control system present at a four-way junction. This ML technique is mainly aimed to replace the existing traffic light control system with artificial intelligence system. Nowadays most cities are equipped with CCTV cameras on the roads and the junctions, the basic idea is to collect the live video from the CCTV cameras and detect the number of vehicles on each lane and feed the data into another machine learning algorithm. according to the data of each lane changes into the light phase of the green signal. This system mainly aims to increase the traffic efficiency by increasing vehicle flow which will reduce waiting time for the vehicles. We are using HOG algorithm for feature extraction. In the implementation of the proposed architecture, we have achieved an accuracy of 86.34% for binary classification and 90.23% for multi-class classification

Design of VLSI Based Power-Saving Streetlight System and Automatic Traffic Controller with Automatic Transfer Switch

The number of road accidents is increasing due to the power outage of the traffic light system and the streetlight system's maintenance fees was too high. Therefore, a proposed solution that includes automatic traffic light controller, energy-saving streetlight controller and automatic energy transfer switch had been designed to overcome these issues. The proposed traffic light is on a four-way junction which has one sensor for each direction. This traffic light controller has 3 different conditions which will act differently when all directions had detected vehicle, or when vehicle is not detected, or when vehicle is detected in some directions. Apart from this, a streetlight controller is also designed to minimize the power usage and the streetlight will only turn on when vehicle is detected. The proposed solution was designed in Quartus 18 via Verilog Hardware Description Language (HDL) and functional simulation is used to verify the proposed circuit is functioning correctly The Verilog HDL files were input to the Synopsys Design Compiler for logical synthesis and IC Compiler for physical synthesis. After both logical and physical synthesis, the design was then analysed in terms of speed, area, and power consumption through the generate report commands. Besides, the proposed energy-saving streetlight was programmed into Field Programmable Gate Array (FPGA) and connected with AC Dimmer Module and LED bulb to measure the power consumption and the reduction of power consumption of the LED bulb. As for the result, the designed chip had a total area of 392.002199 μm^2, positive critical timing slack of 0.1 ns and power consumption of 0.1196 mW. The streetlight was able to achieve a reduction in power consumption by 77.87%.

Solution Structure of NPSL2, A Regulatory Element in the oncomiR-1 RNA

The miR-17 ∼ 92a polycistron, also known as oncomiR-1, is commonly overexpressed in multiple cancers and has several oncogenic properties. OncomiR-1 encodes six constituent microRNAs (miRs), each enzymatically processed with different efficiencies. However, the structural mechanism that regulates this differential processing remains unclear. Chemical probing of oncomiR-1 revealed that the Drosha cleavage sites of pri-miR-92a are sequestered in a four-way junction. NPSL2, an independent stem loop element, is positioned just upstream of pri-miR-92a and sequesters a crucial part of the sequence that constitutes the basal helix of pri-miR-92a. Disruption of the NPSL2 hairpin structure could promote the formation of a pri-miR-92a structure that is primed for processing by Drosha. Thus, NPSL2 is predicted to function as a structural switch, regulating pri-miR-92a processing. Here, we determined the solution structure of NPSL2 using solution NMR spectroscopy. This is the first high-resolution structure of an oncomiR-1 element. NPSL2 adopts a hairpin structure with a large, but highly structured, apical and internal loops. The 10-bp apical loop contains a pH-sensitive A+·C mismatch. Additionally, several adenosines within the apical and internal loops have elevated pKa values. The protonation of these adenosines can stabilize the NPSL2 structure through electrostatic interactions. Our study provides fundamental insights into the secondary and tertiary structure of an important RNA hairpin proposed to regulate miR biogenesis.

Interaction of Proteins with Inverted Repeats and Cruciform Structures in Nucleic Acids.

Cruciforms occur when inverted repeat sequences in double-stranded DNA adopt intra-strand hairpins on opposing strands. Biophysical and molecular studies of these structures confirm their characterization as four-way junctions and have demonstrated that several factors influence their stability, including overall chromatin structure and DNA supercoiling. Here, we review our understanding of processes that influence the formation and stability of cruciforms in genomes, covering the range of sequences shown to have biological significance. It is challenging to accurately sequence repetitive DNA sequences, but recent advances in sequencing methods have deepened understanding about the amounts of inverted repeats in genomes from all forms of life. We highlight that, in the majority of genomes, inverted repeats are present in higher numbers than is expected from a random occurrence. It is, therefore, becoming clear that inverted repeats play important roles in regulating many aspects of DNA metabolism, including replication, gene expression, and recombination. Cruciforms are targets for many architectural and regulatory proteins, including topoisomerases, p53, Rif1, and others. Notably, some of these proteins can induce the formation of cruciform structures when they bind to DNA. Inverted repeat sequences also influence the evolution of genomes, and growing evidence highlights their significance in several human diseases, suggesting that the inverted repeat sequences and/or DNA cruciforms could be useful therapeutic targets in some cases.

The DNA-Binding High-Mobility Group Box Domain of Sox Family Proteins Directly Interacts with RNA In Vitro.

There is a growing body of evidence that a substantial number of protein domains identified as DNA-binding also interact with RNA to regulate biological processes. Several recent studies have revealed that the Sox2 transcription factor binds RNA through its high-mobility group box (HMGB) domain in vitro and in vivo. A high degree of conservation of this domain among members of the Sox family of transcription factors suggests that RNA-binding activity may be a general feature of these proteins. To address this hypothesis, we examined a subset of HMGB domains from human Sox family of proteins for their ability to bind both DNA and RNA in vitro. We observed selective, high-affinity interactions between Sox family HMGB domains and various model RNA elements, including a four-way junction RNA, a hairpin RNA with an internal bulge, G-quadruplex RNA, and a fragment of long noncoding RNA ES2, which is known to directly interact with Sox2. Importantly, the HMGB domains bind these RNA ligands significantly tighter than nonconsensus dsDNA and in some cases with affinities rivaling those of their consensus dsDNA sequences. These data suggest that RNA binding is a conserved feature of the Sox family of transcription factors with the potential to modulate unappreciated biological functions.

The minor groove topology of nucleic acid ligands plays a key role in nuclease resistance and targeting of HMGB1

The architectural DNA‐binding protein High Mobility Group B1 (HMGB1) binds distorted double stranded DNA (e.g. kinked, cruciform) in the nucleus with extremely high affinity to modulate nuclear homeostasis. HMGB1 is a multifunctional protein that also plays a pivotal role outside of the nucleus. It is now clear that HMGB1 can also function as a Damage Molecular Pattern Molecule. In this capacity, HMGB1 can act as a friend or foe to: i)bolster innate immunity to ward off pathogens or ii)exacerbate unwanted immune responses that worsen disease states (e.g. lupus, rheumatoid arthritis). Research shows that cruciform DNA [four‐way junctions (4WJs)] and single strand DNA (ssDNA) can effectively bind/sequester HMGB1. The 4WJs and ssDNA investigated contain natural and phosphorothioate (PS) bonds. PS bonds are introduced to enhance nuclease resistance. Our prior study shows that intramolecular 4WJs composed of natural and PS bonds possess enhanced resistance against the endo‐ and exonucleases. We hypothesize that the increase in nuclease resistance is due to a combination of three factors: i)restricting access to the helical termini (end‐capping), ii)the presence of PS bonds and iii)altered minor groove topology. The objective of this study is to more stringently investigate the potential connection between minor groove topology and nuclease binding. Here, three nucleases are used that cleave double and ssDNA: DNase I, Exonuclease III (Exo III) and bacteriophage T5 Exonuclease (T5 Exo). The nucleases bind within the minor groove to initiate hydrolysis via a non‐specific (DNase I) or specific manner (Exo III and T5 Exo). We hypothesize that nucleases do not bind strongly to nucleic acid substrates with altered (likely wider) minor groove dimensions. As a result, nucleic acids with wider minor grooves tend to have enhanced nuclease resistance. Circular dichroism (CD) studies are used to investigate the minor groove topology of intramolecular 4WJs and ssDNA. The minor groove binder 4’,6‐diamidino‐2‐phenylindole (DAPI) is used to estimate the gross minor groove topology of each nucleic acid ligand. The data clearly shows the DNA substrates with altered minor groove dimensions, as reflected by weakened DAPI binding interactions, possess enhanced nuclease resistance. DNA substrates with enhanced nuclease resistance will be investigated further to target the DNA‐binding cytokine, HMGB1.

MDM2 binds and ubiquitinates PARP1 to enhance DNA replication fork progression.

The MDM2 oncoprotein antagonizes the tumor suppressor p53 by physical interaction and ubiquitination. However, it also sustains the progression of DNA replication forks, even in the absence of functional p53. Here, we show that MDM2 binds, inhibits, ubiquitinates, and destabilizes poly(ADP-ribose) polymerase 1 (PARP1). When cellular MDM2 levels are increased, this leads to accelerated progression of DNA replication forks, much like pharmacological inhibition of PARP1. Conversely, overexpressed PARP1 restores normal fork progression despite elevated MDM2. Strikingly, MDM2 profoundly reduces the frequency of fork reversal, revealed as four-way junctions through electron microscopy. Depletion of RECQ1 or the primase/polymerase (PRIMPOL) reverses the MDM2-mediated acceleration of the nascent DNA elongation rate. MDM2 also increases the occurrence of micronuclei, and it exacerbates camptothecin-induced cell death. In conclusion, high MDM2 levels phenocopy PARP inhibition in modulation of fork restart, representing a potential vulnerability of cancer cells.

Construction of Multiple Logic Circuits Based on Allosteric DNAzymes.

In DNA computing, the implementation of complex and stable logic operations in a universal system is a critical challenge. It is necessary to develop a system with complex logic functions based on a simple mechanism. Here, the strategy to control the secondary structure of assembled DNAzymes’ conserved domain is adopted to regulate the activity of DNAzymes and avoid the generation of four-way junctions, and makes it possible to implement basic logic gates and their cascade circuits in the same system. In addition, the purpose of threshold control achieved by the allosteric secondary structure implements a three-input DNA voter with one-vote veto function. The scalability of the system can be remarkably improved by adjusting the threshold to implement a DNA voter with 2n + 1 inputs. The proposed strategy provides a feasible idea for constructing more complex DNA circuits and a highly integrated computing system.

Finding and cleaving a holliday junction in long double-stranded DNA: a complete resolution reaction trajectory by single-molecule tracking

Resolution of four-way Holliday junctions (HJs) is a critical intermediate step of Homologous Recombination. HJs are processed by abundant junction-resolving endonucleases that introduce two hydrolytic cleavages. Although their catalytic activity and interaction mode with HJ-DNA have been characterized in great detail by biochemical and structural studies, it remains unclear how the endonucleases find their substrate located in kilobase pairs of duplex DNA. Here, we studied the interaction of the T7 endonuclease I with a long dsDNA molecule that has a junction located at its center.

Structural switching/polymorphism by sequential base substitution at quasi-palindromic SNP site (G → A) in LCR of human β-globin gene cluster

The human β-globin gene Locus Control Region (LCR), a dominant regulator of globin gene expression contains five tissue-specific DNase I-hypersensitive sites (HSs). A single nucleotide polymorphism (SNP) (A → G) present in HS4 region of locus control region (LCR), have shown a notable association between the G allele and the occurrence of β-thalassemia. This SNP site exhibiting a hairpin − duplex equilibrium manifested in A → B like DNA transition has previously been reported from this laboratory. Since, DNA is a dynamic and adaptable molecule, so any change of a single base within a primary DNA sequence can produce major biological consequences commonly manifested in genetic disorders such as sickle cell anemia and β-thalassemia. Herein, the differential behavior of sequential single base substitutions G → A on the quasi-palindromic sequence (d-TGGGGGCCCCA; HPG11) has been explored. A combination of native gel electrophoresis, circular dichroism (CD), and UV-thermal denaturation (Tm) techniques have been used to investigate the structural polymorphism associated with various variants of HPG11 i.e. HPG11A2 to HPG11A5. The CD spectra confirmed that all the HPG11 variants exhibit a hairpin − duplex equilibrium. Oligomer concentration dependence on CD spectra has been correlated with A → B DNA conformational transition. However, as revealed in gel electrophoresis, HPG11A2 → A5 exhibit the formation of a tetramolecular structure (four-way junction) at higher oligomer concentration. UV-melting studies also supported the melting of hairpin, duplex and four-way junction structure. This polymorphism pattern may possibly be significant for DNA-protein recognition, in the process of regulation of LCR in the β-globin gene.

Computational investigation of the impact of core sequence on immobile DNA four-way junction structure and dynamics.

Immobile four-way junctions (4WJs) are core structural motifs employed in the design of programmed DNA assemblies. Understanding the impact of sequence on their equilibrium structure and flexibility is important to informing the design of complex DNA architectures. While core junction sequence is known to impact the preferences for the two possible isomeric states that junctions reside in, previous investigations have not quantified these preferences based on molecular-level interactions. Here, we use all-atom molecular dynamics simulations to investigate base-pair level structure and dynamics of four-way junctions, using the canonical Seeman J1 junction as a reference. Comparison of J1 with equivalent single-crossover topologies and isolated nicked duplexes reveal conformational impact of the double-crossover motif. We additionally contrast J1 with a second junction core sequence termed J24, with equal thermodynamic preference for each isomeric configuration. Analyses of the base-pair degrees of freedom for each system, free energy calculations, and reduced-coordinate sampling of the 4WJ isomers reveal the significant impact base sequence has on local structure, isomer bias, and global junction dynamics.

Self-assembled RNA nanocarrier-mediated chemotherapy combined with molecular targeting in the treatment of esophageal squamous cell carcinoma

BackgroundEsophageal cancer is the fifth most common cancer affecting men in China. The primary treatment options are surgery and traditional radio-chemotherapy; no effective targeted therapy exists yet. Self-assembled RNA nanocarriers are highly stable, easily functionally modified, and have weak off-tumor targeting effects. Thus, they are among the most preferred carriers for mediating the targeted delivery of anti-tumor drugs. miR-375 was found to be significantly down-regulated in esophageal squamous cell carcinoma (ESCC) tissues and its overexpression effectively inhibits the proliferation, migration, and invasion of ESCC cells. Moreover, epidermal growth factor receptor (EGFR) was overexpressed in ESCC cells, and accumulation of RNA nanoparticles in ESCC tumors was enhanced by EGFR-specific aptamer (EGFRapt) modification.ResultsHerein, a novel four-way junction RNA nanocarrier, 4WJ-EGFRapt-miR-375-PTX simultaneously loaded with miR-375, PTX and decorated with EGFRapt, was developed. In vitro analysis demonstrated that 4WJ-EGFRapt-miR-375-PTX possesses strong thermal and pH stabilities. EGFRapt decoration facilitated tumor cell endocytosis and promoted deep penetration into 3D-ESCC spheroids. Xenograft mouse model for ESCC confirmed that 4WJ-EGFRapt-miR-375-PTX was selectively distributed in tumor sites via EGFRapt-mediating active targeting and targeted co-delivery of miR-375 and PTX exhibited more effective therapeutic efficacy with low systemic toxicity.ConclusionThis strategy may provide a practical approach for targeted therapy of ESCC.Graphical

Dual Catalytic Hairpin Assembly-Based Automatic Molecule Machine for Amplified Detection of Auxin Response Factor-Targeted MicroRNA-160.

MicroRNA160 plays a crucial role in plant development by negatively regulating the auxin response factors (ARFs). In this manuscript, we design an automatic molecule machine (AMM) based on the dual catalytic hairpin assembly (D-CHA) strategy for the signal amplification detection of miRNA160. The detection system contains four hairpin-shaped DNA probes (HP1, HP2, HP3, and HP4). For HP1, the loop is designed to be complementary to miRNA160. A fragment of DNA with the same sequences as miRNA160 is separated into two pieces that are connected at the 3′ end of HP2 and 5′ end of HP3, respectively. In the presence of the target, four HPs are successively dissolved by the first catalytic hairpin assembly (CHA1), forming a four-way DNA junction (F-DJ) that enables the rearrangement of separated DNA fragments at the end of HP2 and HP3 and serving as an integrated target analogue for initiating the second CHA reaction, generating an enhanced fluorescence signal. Assay experiments demonstrate that D-CHA has a better performance compared with traditional CHA, achieving the detection limit as low as 10 pM for miRNA160 as deduced from its corresponding DNA surrogates. Moreover, non-target miRNAs, as well as single-base mutation targets, can be detected. Overall, the D-CHA strategy provides a competitive method for plant miRNAs detection.

Rapid, multiplexed, and nucleic acid amplification-free detection of SARS-CoV-2 RNA using an electrochemical biosensor

Considering the worldwide health crisis associated with highly contagious severe respiratory disease of COVID-19 outbreak, the development of multiplexed, simple and rapid diagnostic platforms to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is in high demand. Here, a nucleic acid amplification-free electrochemical biosensor based on four-way junction (4-WJ) hybridization is presented for the detection of SARS-CoV-2. To form a 4-WJ structure, a Universal DNA-Hairpin (UDH) probe is hybridized with two adaptor strands and a SARS-CoV-2 RNA target. One of the adaptor strands is functionalized with a redox mediator that can be detected using an electrochemical biosensor. The biosensor could simultaneously detect 5.0 and 6.8 ag/μL of S and Orf1ab genes, respectively, within 1 h. The biosensor was evaluated with 21 clinical samples (16 positive and 5 negative). The results revealed a satisfactory agreement with qRT-PCR. In conclusion, this biosensor has the potential to be used as an on-site, real-time diagnostic test for COVID-19.

Functional Nucleic Acids for Pathogenic Bacteria Detection.

Pathogens have long presented a significant threat to human lives, and hence the rapid detection of infectious pathogens is vital for improving human health. Current detection methods lack the means to detect infectious pathogens in a simple, rapid, and reliable manner at the time and point of need. Functional nucleic acids (FNAs) have the potential to overcome these limitations by acting as key components for point-of-care (POC) biosensors due to their distinctive advantages that include high binding affinities and specificities, excellent chemical stability, ease of synthesis and modification, and compatibility with a variety of signal-amplification and signal-transduction mechanisms.This Account summarizes the work completed in our groups toward developing FNA-based biosensors for detecting bacteria. In vitro selection has led to the isolation of many RNA-cleaving fluorogenic DNAzymes (RFDs) and DNA aptamers that can recognize infectious pathogens, including Escherichia coli, Clostridium difficile, Helicobacter pylori, and Legionella pneumophila. In most cases, a "many-against-many" approach was employed using a DNA library against a crude cellular mixture of an infectious pathogen containing diverse biomarkers as the target to isolate RFDs, with combined counter and positive selections ensuring high specificity toward the desired target. This procedure allows for the isolation of pathogen-specific FNAs without first identifying a suitable biomarker. Multiple target-specific DNA aptamers, including anti-glutamate dehydrogenase (GDH) circular aptamers, anti-degraded toxin B aptamers, and anti-RNase HII aptamers, have also been isolated for the detection of bacteria such as Clostridium difficile. The isolated FNAs have been integrated into fluorescent, colorimetric, and electrochemical biosensors using various signal transduction mechanisms. Both simple-to-use paper-based analytical devices and hand-held electrical devices with integrated FNAs have been developed for POC applications. In addition, signal-amplification strategies, including DNA catenane enabled rolling circle amplification (RCA), DNAzyme feedback RCA, and an all-DNA amplification system using a four-way junction and catalytic hairpin assembly (CHA), have been designed and applied to these systems to further increase their detection sensitivity. The use of these FNA-based biosensors to detect pathogens directly in clinical samples, such as urine, blood, and stool, has now been demonstrated with an outstanding sensitivity of as low as 10 cells per milliliter, highlighting the tremendous potential of using FNA-based sensors in clinical applications. We further describe strategies to overcome the challenges of using FNA-based biosensors in clinical applications, including strategies to improve the stability of FNAs in biological samples and prevent their nonspecific degradation from nucleases and strategies to deal with issues such as signal loss caused by nonspecific binding and biofouling. Finally, the remaining roadblocks for employing FNA-based biosensors in clinical applications are discussed.

Effect of external HMI for automated vehicles (AVs) on drivers’ ability to infer the AV motion intention: A field experiment

A field experiment was conducted aiming to shed light on how drivers negotiate anambiguous traffic situation when encountering an autonomous vehicle (AV) in the presence of a yielding intention signal (AV with eHMI) or not (AV without eHMI). A traffic conflict scenario was created with two opposing vehicles instructed to perform a left turn at a four-way junction, at the same point in time. Forty participants were assigned to two groups encountering either an AV with eHMI or an AV without eHMI. To check for equivalence across the two groups, both groups also encountered a conventional vehicle (CV).Results showed that the two groups performed similarly during encounters with a CV. During encounters with AV, however, participants provided with the eHMI maintained higher speed before the AV and finished their maneuver quicker than when they were not provided with eHMI. Additionally participants provided with eHMI rated higher their ability to infer the AV intention before coming to a full-stop than those provided without eHMI. The above findings indicate that the presence of eHMI on AV can accelerate drivers’ inferences about yielding intention of an AV, and have the potential to optimize AV-driver interaction in ambiguous traffic situations.

Overexpression of OsHMGB707, a High Mobility Group Protein, Enhances Rice Drought Tolerance by Promoting Stress-Related Gene Expression.

Drought stress adversely affects crop growth and productivity worldwide. In response, plants have evolved several strategies in which numerous genes are induced to counter stress. High mobility group (HMG) proteins are the second most abundant family of chromosomal proteins. They play a crucial role in gene transcriptional regulation by modulating the chromatin/DNA structure. In this study, we isolated a novel HMG gene, OsHMGB707, one of the candidate genes localized in the quantitative trait loci (QTL) interval of rice drought tolerance, and examined its function on rice stress tolerance. The expression of OsHMGB707 was up-regulated by dehydration and high salt treatment. Its overexpression significantly enhanced drought tolerance in transgenic rice plants, whereas its knockdown through RNA interference (RNAi) did not affect the drought tolerance of the transgenic rice plants. Notably, OsHMGB707-GFP is localized in the cell nucleus, and OsHMGB707 is protein-bound to the synthetic four-way junction DNA. Several genes were up-regulated in OsHMGB707-overexpression (OE) rice lines compared to the wild-type rice varieties. Some of the genes encode stress-related proteins (e.g., DREB transcription factors, heat shock protein 20, and heat shock protein DnaJ). In summary, OsHMGB707 encodes a stress-responsive high mobility group protein and regulates rice drought tolerance by promoting the expression of stress-related genes.

Three-dimensional structure of a flavivirus dumbbell RNA reveals molecular details of an RNA regulator of replication.

Mosquito-borne flaviviruses (MBFVs) including dengue, West Nile, yellow fever, and Zika viruses have an RNA genome encoding one open reading frame flanked by 5′ and 3′ untranslated regions (UTRs). The 3′ UTRs of MBFVs contain regions of high sequence conservation in structured RNA elements known as dumbbells (DBs). DBs regulate translation and replication of the viral RNA genome, functions proposed to depend on the formation of an RNA pseudoknot. To understand how DB structure provides this function, we solved the x-ray crystal structure of the Donggang virus DB to 2.1Å resolution and used structural modeling to reveal the details of its three-dimensional fold. The structure confirmed the predicted pseudoknot and molecular modeling revealed how conserved sequences form a four-way junction that appears to stabilize the pseudoknot. Single-molecule FRET suggests that the DB pseudoknot is a stable element that can regulate the switch between translation and replication during the viral lifecycle by modulating long-range RNA conformational changes.