Targeting the NTF2-like domain of G3BP1: Novel modulators of intracellular granule dynamics.

Stress granule (SG) is a temporary cellular structure that plays a crucial role in the regulation of mRNA and protein sequestration during various cellular stress conditions. SG enables cells to cope with stress more effectively, conserving vital energy and resources. Focusing on the NTF2-like domain of G3BP1, a key protein in SG dynamics, we explore to identify and characterize novel small molecules involved in SG modulation without external stressors. Through in silico molecular docking approach to simulate the interaction between various compounds and the NTF2-like domain of G3BP1, we identified three compounds as potential candidates that could bind to the NTF2-like domain of G3BP1. Subsequent immunofluorescence experiments demonstrated that these compounds induce the formation of SG-like, G3BP1-positive granules. Importantly, the granule formation by these compounds occurs independent from the phosphorylation of eIF2α, a common mechanism in SG formation, suggesting that it might offer a new strategy for influencing SG dynamics implicated in various diseases.

Ribonucleoprotein Granules: Between Stress and Transposable Elements.

Transposable elements (TEs) are DNA sequences that can transpose and replicate within the genome, leading to genetic changes that affect various aspects of host biology. Evolutionarily, hosts have also developed molecular mechanisms to suppress TEs at the transcriptional and post-transcriptional levels. Recent studies suggest that stress-induced formation of ribonucleoprotein (RNP) granules, including stress granule (SG) and processing body (P-body), can play a role in the sequestration of TEs to prevent transposition, suggesting an additional layer of the regulatory mechanism for TEs. RNP granules have been shown to contain factors involved in RNA regulation, including mRNA decay enzymes, RNA-binding proteins, and noncoding RNAs, which could potentially contribute to the regulation of TEs. Therefore, understanding the interplay between TEs and RNP granules is crucial for elucidating the mechanisms for maintaining genomic stability and controlling gene expression. In this review, we provide a brief overview of the current knowledge regarding the interplay between TEs and RNP granules, proposing RNP granules as a novel layer of the regulatory mechanism for TEs during stress.

A Fluorescent and Colorimetric Chemosensor Detecting Pd2+ Based on Chalcone Structure with Triphenylamine.

A fluorometric and colorimetric chemosensor DiPP ((E)-3-(4-(diphenylamino)phenyl)-1-(pyridin-2-yl)prop-2-en-1-one) based on chalcone structure with a triphenylamine group was synthesized. Sensor DiPP detected Pd2+ with fluorescence turn-off and via colorimetry variation of yellow to purple. The binding ratio of DiPP to Pd2+ turned out to be 1 : 1. Detection limits for Pd2+ by DiPP were analyzed to be 0.67 µM and 0.80 µM through the fluorescent and colorimetric methods. Additionally, the fluorescent and colorimetric test strips were applied for probing Pd2+ and displayed that DiPP could obviously discriminate Pd2+ from other metals. The binding feature of DiPP to Pd2+ was presented by ESI-mass, Job plot, NMR titration, ESI-mass, and DFT calculations.

Ongoing endeavors to detect mobilization of transposable elements.

Transposable elements (TEs) are DNA sequences capable of mobilization from one location to another in the genome. Since the discovery of 'Dissociation (Dc) locus' by Barbara McClintock in maize (1), mounting evidence in the era of genomics indicates that a significant fraction of most eukaryotic genomes is composed of TE sequences, involving in various aspects of biological processes such as development, physiology, diseases and evolution. Although technical advances in genomics have discovered numerous functional impacts of TE across species, our understanding of TEs is still ongoing process due to challenges resulted from complexity and abundance of TEs in the genome. In this mini-review, we briefly summarize biology of TEs and their impacts on the host genome, emphasizing importance of understanding TE landscape in the genome. Then, we introduce recent endeavors especially in vivo retrotransposition assays and long read sequencing technology for identifying de novo insertions/TE polymorphism, which will broaden our knowledge of extraordinary relationship between genomic cohabitants and their host. [BMB Reports 2022; 55(7): 305-315].

Rapamycin Promotes ROS-Mediated Cell Death via Functional Inhibition of xCT Expression in Melanoma Under γ-Irradiation.

Although many cancer patients are administered radiotherapy for their treatment, the interaction between tumor cells and macrophages in the tumor microenvironment attenuates the curative effects of radiotherapy. The enhanced activation of mTOR signaling in the tumors promotes tumor radioresistance. In this study, the effects of rapamycin on the interaction between tumor cells and macrophages were investigated. Rapamycin and 3BDO were used to regulate the mTOR pathway. In vitro, tumor cells cocultured with macrophages in the presence of each drug under normoxic or hypoxic conditions were irradiated with γ-rays. In vivo, mice were irradiated with γ-radiation after injection with DMSO, rapamycin and 3BDO into tumoral regions. Rapamycin reduced the secretion of IL-4 in tumor cells as well as YM1 in macrophages. Mouse recombinant YM1 decreased the enhanced level of ROS and the colocalized proportion of both xCT and EEA1 in irradiated tumor cells. Human recombinant YKL39 also induced results similar to those of YM1. Moreover, the colocalized proportion of both xCT and LC3 in tumor tissues was elevated by the injection of rapamycin into tumoral regions. Overall, the suppression of mTOR signaling in the tumor microenvironment might be useful for the improvement of tumor radioresistance.

A novel role of Hippo-Yap/TAZ signaling pathway in lymphatic vascular development.

The lymphatic vasculature plays important role in regulating fluid homeostasis, intestinal lipid absorption, and immune surveillance in humans. Malfunction of lymphatic vasculature leads to several human diseases. Understanding the fundamental mechanism in lymphatic vascular development not only expand our knowledge, but also provide a new therapeutic insight. Recently, Hippo-YAP/TAZ signaling pathway, a key mechanism of organ size and tissue homeostasis, has emerged as a critical player that regulate lymphatic specification, sprouting, and maturation. In this review, we discuss the mechanistic regulation and pathophysiological significant of Hippo pathway in lymphatic vascular development. [BMB Reports 2021; 54(6): 285-294].

PCM1 is necessary for focal ciliary integrity and is a candidate for severe schizophrenia.

The neuronal primary cilium and centriolar satellites have functions in neurogenesis, but little is known about their roles in the postnatal brain. We show that ablation of pericentriolar material 1 in the mouse leads to progressive ciliary, anatomical, psychomotor, and cognitive abnormalities. RNAseq reveals changes in amine- and G-protein coupled receptor pathways. The physiological relevance of this phenotype is supported by decreased available dopamine D2 receptor (D2R) levels and the failure of antipsychotic drugs to rescue adult behavioral defects. Immunoprecipitations show an association with Pcm1 and D2Rs. Finally, we sequence PCM1 in two human cohorts with severe schizophrenia. Systematic modeling of all discovered rare alleles by zebrafish in vivo complementation reveals an enrichment for pathogenic alleles. Our data emphasize a role for the pericentriolar material in the postnatal brain, with progressive degenerative ciliary and behavioral phenotypes; and they support a contributory role for PCM1 in some individuals diagnosed with schizophrenia.

Lysophosphatidylcholine Enhances Bactericidal Activity by Promoting Phagosome Maturation via the Activation of the NF-κB Pathway during Salmonella Infection in Mouse Macrophages.

Salmonella enterica serovar Typhimurium (S. Typhimurium) is a facultative intracellular pathogen that causes salmonellosis and mortality worldwide. S. Typhimurium infects macrophages and survives within phagosomes by avoiding the phagosome-lysosome fusion system. Phagosomes sequentially acquire different Rab GTPases during maturation and eventually fuse with acidic lysosomes. Lysophosphatidylcholine (LPC) is a bioactive lipid that is associated with the generation of chemoattractants and reactive oxygen species (ROS). In our previous study, LPC controlled the intracellular growth of Mycobacterium tuberculosis by promoting phagosome maturation. In this study, to verify whether LPC enhances phagosome maturation and regulates the intracellular growth of S. Typhimurium, macrophages were infected with S. Typhimurium. LPC decreased the intracellular bacterial burden, but it did not induce cytotoxicity in S. Typhimuriuminfected cells. In addition, combined administration of LPC and antibiotic significantly reduced the bacterial burden in the spleen and the liver. The ratios of the colocalization of intracellular S. Typhimurium with phagosome maturation markers, such as early endosome antigen 1 (EEA1) and lysosome-associated membrane protein 1 (LAMP-1), were significantly increased in LPC-treated cells. The expression level of cleaved cathepsin D was rapidly increased in LPCtreated cells during S. Typhimurium infection. Treatment with LPC enhanced ROS production, but it did not affect nitric oxide production in S. Typhimurium-infected cells. LPC also rapidly triggered the phosphorylation of IκBα during S. Typhimurium infection. These results suggest that LPC can improve phagosome maturation via ROS-induced activation of NF-κB pathway and thus may be developed as a therapeutic agent to control S. Typhimurium growth.

Knockout of SETDB1 gene using the CRISPR/cas-9 system increases migration and transforming activities via complex regulations of E-cadherin, ß-catenin, STAT3, and Akt.

SETDB1 HMTase participates in various cellular processes via epigenetic transcriptional regulation. SETDB1 expression is downregulated by anticancer drug treatment in cancer cells, but we still need to verify the functional significance on SETDB1 downregulation. CRISPR/cas9 is a useful technology for doing a knockout (KO) of a target gene. It is widely used to examine the function of genes. In this study, we prepared SETDB1-KO from A549 human lung cancer cells using the CRISPR/Cas9 system, and we compared molecular changes between the A549 cells and the SETDB1-KO cells. The SETDB1-KO cell proliferation rate was slightly decreased as compared to the A549 cells, but there was no large difference in sensitivity with doxorubicin treatment. Instead, the migration activity and transforming activity were dramatically increased in SETDB-KO cells. Using a western blot analysis and an immunostaining experiment, we confirmed that SETDB1-KO downregulates the expression of E-cadherin and ß-catenin. A qPCR and an RT-PCR analysis suggested that SETDB1 transcriptionally regulates E-cadherin and ß-catenin. Moreover, E-cadherin expression was also detected in the cytoplasmic region of SETDB1-KO cells, indicating that functional localization of E-cadherin might be changed in SETDB1-KO cells. On the other hand, total levels of STAT3 and Akt were increased in the SETDB1-KO cells, but activation of STAT3 (pSTAT3) was not induced in doxorubicin-treated SETDB1-KO cells. SETDB1 overexpression into SETDB1-KO cells restores the expression of E-cadherin, ß-catenin, STAT3, and Akt, suggesting that those proteins are tightly regulated by SETDB1. Collectively, we suggest that complex regulations on E-cadherin, ß-catenin, STAT3, and Akt are correlated with the increased migration and transforming activity of SETDB1-KO cells.

The optimization of surface morphology of Au nanoparticles on WO3 nanoflakes for plasmonic photoanode.

Among many candidates for photoanode materials of photoelectrochemical (PEC) cell, nanostructured tungsten trioxide (WO3) is regarded as one of the most promising materials due to its superior electrical properties and adequate bandgap (∼2.8 eV) and band edge position. WO3 nanoflakes (WO3 NFs), which have merits on its high surface area and crystallinity, have been actively studied for this manner but solar-to-hydrogen efficiency of WO3 NFs based photoanode is still not sufficient both in light absorption and charge separation. Plasmon-induced enhancement using Au nanoparticles is excellent approach for both the efficiency of light absorption and charge separation of WO3. However, it still needs optimization on its amount, shape, coverage, and etc. Here, we synthesized WO3 NFs by solvothermal growth and decorated gold nanoparticles on these nanoflakes by e-beam evaporation and rapid thermal annealing process in a row. By this process, a large-area AuNPs/WO3 nanocomposite structure with various size, interparticle distance, and coverage of AuNPs were fabricated. These AuNPs/WO3 NFs type photoanode achieve high light absorption both in UV and visible range and consequently higher photocurrent density. The optimized AuNPs/WO3 nanocomposite photoanode exhibits 1.01 mA cm-2 of photocurrent density, which is increased to 19.8% compared with bare WO3 nanoflakes. Field emission-scanning electron microscope, x-ray diffraction, UV-vis spectrometer analysis were measured to analyze the morphology and crystallinity and relationship between structure and PEC performance.

Pathogenic variants in E3 ubiquitin ligase RLIM/RNF12 lead to a syndromic X-linked intellectual disability and behavior disorder.

RLIM, also known as RNF12, is an X-linked E3 ubiquitin ligase acting as a negative regulator of LIM-domain containing transcription factors and participates in X-chromosome inactivation (XCI) in mice. We report the genetic and clinical findings of 84 individuals from nine unrelated families, eight of whom who have pathogenic variants in RLIM (RING finger LIM domain-interacting protein). A total of 40 affected males have X-linked intellectual disability (XLID) and variable behavioral anomalies with or without congenital malformations. In contrast, 44 heterozygous female carriers have normal cognition and behavior, but eight showed mild physical features. All RLIM variants identified are missense changes co-segregating with the phenotype and predicted to affect protein function. Eight of the nine altered amino acids are conserved and lie either within a domain essential for binding interacting proteins or in the C-terminal RING finger catalytic domain. In vitro experiments revealed that these amino acid changes in the RLIM RING finger impaired RLIM ubiquitin ligase activity. In vivo experiments in rlim mutant zebrafish showed that wild type RLIM rescued the zebrafish rlim phenotype, whereas the patient-specific missense RLIM variants failed to rescue the phenotype and thus represent likely severe loss-of-function mutations. In summary, we identified a spectrum of RLIM missense variants causing syndromic XLID and affecting the ubiquitin ligase activity of RLIM, suggesting that enzymatic activity of RLIM is required for normal development, cognition and behavior.

A Robust Transposon-Endogenizing Response from Germline Stem Cells.

The heavy occupancy of transposons in the genome implies that existing organisms have survived from multiple, independent rounds of transposon invasions. However, how and which host cell types survive the initial wave of transposon invasion remain unclear. We show that the germline stem cells can initiate a robust adaptive response that rapidly endogenizes invading P element transposons by activating the DNA damage checkpoint and piRNA production. We find that temperature modulates the P element activity in germline stem cells, establishing a powerful tool to trigger transposon hyper-activation. Facing vigorous invasion, Drosophila first shut down oogenesis and induce selective apoptosis. Interestingly, a robust adaptive response occurs in ovarian stem cells through activation of the DNA damage checkpoint. Within 4 days, the hosts amplify P element-silencing piRNAs, repair DNA damage, subdue the transposon, and reinitiate oogenesis. We propose that this robust adaptive response can bestow upon organisms the ability to survive recurrent transposon invasions throughout evolution.

Hijacking Oogenesis Enables Massive Propagation of LINE and Retroviral Transposons.

Although animals have evolved multiple mechanisms to suppress transposons, "leaky" mobilizations that cause mutations and diseases still occur. This suggests that transposons employ specific tactics to accomplish robust propagation. By directly tracking mobilization, we show that, during a short and specific time window of oogenesis, retrotransposons achieve massive amplification via a cell-type-specific targeting strategy. Retrotransposons rarely mobilize in undifferentiated germline stem cells. However, as oogenesis proceeds, they utilize supporting nurse cells-which are highly polyploid and eventually undergo apoptosis-as factories to massively manufacture invading products. Moreover, retrotransposons rarely integrate into nurse cells themselves but, instead, via microtubule-mediated transport, they preferentially target the DNA of the interconnected oocytes. Blocking microtubule-dependent intercellular transport from nurse cells significantly alleviates damage to the oocyte genome. Our data reveal that parasitic genomic elements can efficiently hijack a host developmental process to propagate robustly, thereby driving evolutionary change and causing disease.

An Omnidirectionally Stretchable Photodetector Based on Organic-Inorganic Heterojunctions.

Omnidirectionally stretchable photodetectors are limited by difficulties in designing material and fabrication processes that enable stretchability in multiaxial directions. Here, we propose a new approach involving an organic-inorganic p-n heterojunction photodetector comprised of free-standing ZnO nanorods grown on a poly(3,4-ethylenedioxythiophene)-polystyrene sulfonate transport layer coated on a three-dimensional micropatterned stretchable substrate containing bumps and valleys. This structure allows for efficient absorption of stretching strain. This approach allows the device to accommodate large tensile strain in all of the directions. The device behaves as a photogated p-n heterojunction photodetector in which current modulation was obtained by sensing the mechanisms that rely on photovoltage and photogating effects. The device exhibits a high photoresponse to UV light and reliable electrical performance under applied stretching in uniaxial and omniaxial directions. Furthermore, the device can be easily and conformally attached to a human wrist. This allowed us to investigate the response of the device to UV light during human activity.

Broadband light-absorption InGaN photoanode assisted by imprint patterning and ZnO nanowire growth for energy conversion.

In this research, an InGaN-based photoanode with a broadband light-absorption range from ultraviolet to green, patterned by imprint lithography and branched by ZnO nanowires, has been applied to water splitting. Over the solar spectrum range, the absorbance increases due to the scattering effect of the micro-structure compared to that of flat surface InGaN, which reaches a maximum of over 90% at 380 nm as ZnO nanowires are further employed in this novel photoanode. Consequently, the induced photocurrent density of the InGaN photoanode with a domelike structure and ZnO nanowires on the surface shows a remarkable enhancement of seven times that of the one with a flat surface. Further investigation indicates the wet-etching process for defect removal has an essential impact on photocurrent efficiency. This design demonstrates an innovative approach for water splitting.

Splicing-independent loading of TREX on nascent RNA is required for efficient expression of dual-strand piRNA clusters in Drosophila.

The conserved THO/TREX (transcription/export) complex is critical for pre-mRNA processing and mRNA nuclear export. In metazoa, TREX is loaded on nascent RNA transcribed by RNA polymerase II in a splicing-dependent fashion; however, how TREX functions is poorly understood. Here we show that Thoc5 and other TREX components are essential for the biogenesis of piRNA, a distinct class of small noncoding RNAs that control expression of transposable elements (TEs) in the Drosophila germline. Mutations in TREX lead to defects in piRNA biogenesis, resulting in derepression of multiple TE families, gametogenesis defects, and sterility. TREX components are enriched on piRNA precursors transcribed from dual-strand piRNA clusters and colocalize in distinct nuclear foci that overlap with sites of piRNA transcription. The localization of TREX in nuclear foci and its loading on piRNA precursor transcripts depend on Cutoff, a protein associated with chromatin of piRNA clusters. Finally, we show that TREX is required for accumulation of nascent piRNA precursors. Our study reveals a novel splicing-independent mechanism for TREX loading on nascent RNA and its importance in piRNA biogenesis.

InGaN-based photoanode with ZnO nanowires for water splitting.

The water splitting properties of InGaN photoanodes equipped with ZnO nanowires were examined in this study. Over the solar spectrum range, the absorbance exhibited a remarkable increase due to the enhanced light absorption caused by the ZnO nanowires. By varying the ZnO nanowires length, the photo-to-current density of photoanodes was increased from 0.017 to 0.205 mA/cm2 at 1.23 V versus reversible hydrogen electrode. Consequently, the incident-photon-to-current efficiency was increased by a factor of 5.5 as the ZnO nanowires growth time increased from 2 to 4 h. The results of this research demonstrate the importance of light absorbance and the surface reaction sites of photoanodes on energy harvesting.

TAF1 Variants Are Associated with Dysmorphic Features, Intellectual Disability, and Neurological Manifestations.

We describe an X-linked genetic syndrome associated with mutations in TAF1 and manifesting with global developmental delay, intellectual disability (ID), characteristic facial dysmorphology, generalized hypotonia, and variable neurologic features, all in male individuals. Simultaneous studies using diverse strategies led to the identification of nine families with overlapping clinical presentations and affected by de novo or maternally inherited single-nucleotide changes. Two additional families harboring large duplications involving TAF1 were also found to share phenotypic overlap with the probands harboring single-nucleotide changes, but they also demonstrated a severe neurodegeneration phenotype. Functional analysis with RNA-seq for one of the families suggested that the phenotype is associated with downregulation of a set of genes notably enriched with genes regulated by E-box proteins. In addition, knockdown and mutant studies of this gene in zebrafish have shown a quantifiable, albeit small, effect on a neuronal phenotype. Our results suggest that mutations in TAF1 play a critical role in the development of this X-linked ID syndrome.

A human laterality disorder caused by a homozygous deleterious mutation in MMP21.

BACKGROUND: Laterality in the vertebrate embryo is determined by left-right asymmetric gene expression driven by the flow of extraembryonic fluid across the embryonic node. Defects in these processes cause heterotaxy, the abnormal formation and arrangement of visceral organs that can range from complete inversion of symmetry to the selective misarrangement of organs. However, our understanding of the genetic causality for laterality defects in human beings remains relatively limited. METHODS: We performed whole exome sequencing in a consanguineous family with heterotaxia. To interrogate the pathogenic potential of the discovered variant, we used an in vivo system in which the potential of the candidate gene to induce L-R asymmetry was tested by transient suppression and CRISPR/Cas9-induced deletions. We also used in vitro assays to test a possible link between our exome-derived candidate and Notch signaling. RESULTS: We identified a homozygous 2 bp deletion in MMP21, encoding matrix metalloproteinase-21, as the sole coding mutation that segregated with the phenotype. Transient suppression or CRISPR/Cas9-mediated deletion of mmp21 in zebrafish embryos induced cardiac looping defects, with concomitant disruption of laterality markers in the lateral plate mesoderm and disrupted notch signalling in vitro and in vivo. CONCLUSIONS: Our data implicate loss of MMP21 as a cause of heterotaxy in humans with concomitant defects in Notch signaling. In support of this finding, a homozygous missense mutation in MMP21 was identified previously in mice with N-Ethyl-N-Nitrosourea (ENU)-induced heterotaxy. Taken together, these observations suggest a role of matrix metalloproteinases in the establishment of asymmetric organ development, likely through the regulation of morphogenetic signals.

Development of biomarker for detecting silver nanoparticles exposure using a GAL4 enhancer trap screening in Drosophila.

Silver nanoparticles (AgNPs) have been widely used in commercial goods ranging from medical devices to home appliances. Their widespread application increase the risk related to their potential toxicity. Although several studies showed their acute hazardous effects on living animals, our understanding of chronic effects of AgNPs exposed by the environment we encounter in our everyday lives is still very limited. This is partly because of the lack of versatile animal model system for studying AgNPs effects on terrestrial animals including human. In this study, we used Drosophila model to study AgNPs toxicity in terrestrial animals, and found that long-term exposure of AgNPs, but not Ag ions, at low level (0.1 and 1µg/mL) significantly shortened the lifespan. By taking advantage of the power of Drosophila genetics, we also isolated a GAL4 enhancer trap line called M95, in which the expression of GAL4 is up-regulated in response to ingestion of AgNPs at concentrations as low as 0.1µg/mL. Interestingly M95 flies showed significantly increased tolerance to both AgNPs treatment and dry starvation probably due to up-regulation of JNK signaling. These findings suggest not only that M95 may be a very useful biomarker of AgNPs because of its high sensitivity and tolerance to AgNPs, but also that Drosophila may be a versatile terrestrial invertebrate model for studying the effects of AgNPs on human health.