G-quadruplex DNA contributes to RNA polymerase II-mediated 3D chromatin architecture.

High-order chromatin organization plays an important role in biological processes and disease development. Previous studies revealed a widespread occurrence of guanine quadruplex (G4) structures in the human genome, with enrichment in gene regulatory regions, especially in promoters. However, it remains unclear whether G4 structures contribute to RNA polymerase II (RNAPII)-mediated long-range DNA interactions and transcription activity. In this study, we conducted an intuitive overlapping analysis of previously published RNAPII ChIA-PET (chromatin interaction analysis with paired-end tag) and BG4 ChIP-seq (chromatin immunoprecipitation followed by sequencing using a G4 structure-specific antibody) data. We observed a strong positive correlation between RNAPII-linked DNA loops and G4 structures in chromatin. Additionally, our RNAPII HiChIP-seq (in situ Hi-C followed by ChIP-seq) results showed that treatment of HepG2 cells with pyridostatin (PDS), a small-molecule G4-binding ligand, could diminish RNAPII-linked long-range DNA contacts, with more pronounced diminutions being observed for those contacts involving G4 structure loci. RNA sequencing data revealed that PDS treatment modulates the expression of not only genes with G4 structures in their promoters, but also those with promoters being connected with distal G4s through RNAPII-linked long-range DNA interactions. Together, our data substantiate the function of DNA G4s in RNAPII-associated DNA looping and transcription regulation.

Backbone cyclization of Salmonella typhimurium diaminopropionate ammonia-lyase to enhance the activity and stability.

Diaminopropionate ammonia-lyase transforms D and L isomers of 2,3-diaminopropionate to pyruvate and ammonia. It catalyzes D- and l-serine less effectively. L-2,3-diaminopropionate is a precursor in the biosynthesis of oxalyl diaminopropionate as a neurotoxin in certain legume species. In this work, we cyclized the diaminopropionate ammonia-lyase from Salmonella typhimurium in vitro using the redox-responsive split intein, and identified that backbone cyclization afforded the enzyme with the improved activity, thermal stability and resistance to the exopeptidase proteolysis, different from effects of the incorporated sequence recognized by tobacco vein mottling virus protease at C-terminus. Using analyses of three fluorescent dyes including 8-anilino-1-naphthalenesulfonic acid, N-phenyl-1-naphthylamine, and thioflavin T, the same amounts of the cyclic protein displayed less fluorescence than those of the linear protein upon the heat treatment. The cyclic enzyme displayed the enhanced activity in Escherichia coli cells using the designed novel reporter. In this system, d-serine was added to the culture and transported into the cytoplasm. It was transformed by pre-overexpression of the diaminopropionate ammonia-lyase, and untransformed d-serine was oxidized by the coproduced human d-amino acid oxidase to generate hydrogen peroxide. This oxidant is monitored by the HyPer indicator. The current results presented that the cyclized enzyme could be applied as a better candidate to block the neurotoxin biosynthesis in certain plant species.

Chemomechanics of Rechargeable Batteries: Status, Theories, and Perspectives.

Chemomechanics is an old subject, yet its importance has been revived in rechargeable batteries where the mechanical energy and damage associated with redox reactions can significantly affect both the thermodynamics and rates of key electrochemical processes. Thanks to the push for clean energy and advances in characterization capabilities, significant research efforts in the last two decades have brought about a leap forward in understanding the intricate chemomechanical interactions regulating battery performance. Going forward, it is necessary to consolidate scattered ideas in the literature into a structured framework for future efforts across multidisciplinary fields. This review sets out to distill and structure what the authors consider to be significant recent developments on the study of chemomechanics of rechargeable batteries in a concise and accessible format to the audiences of different backgrounds in electrochemistry, materials, and mechanics. Importantly, we review the significance of chemomechanics in the context of battery performance, as well as its mechanistic understanding by combining electrochemical, materials, and mechanical perspectives. We discuss the coupling between the elements of electrochemistry and mechanics, key experimental and modeling tools from the small to large scales, and design considerations. Lastly, we provide our perspective on ongoing challenges and opportunities ranging from quantifying mechanical degradation in batteries to manufacturing battery materials and developing cyclic protocols to improve the mechanical resilience.

Non-Fluorinated Ethers to Mitigate Electrode Surface Reactivity in High-Voltage NCM811-Li Batteries.

Lithium (Li) metal batteries (LMBs) with nickel (Ni)-rich layered oxide cathodes exhibit twice the energy density of conventional Li-ion batteries. However, their lifespan is limited by severe side reactions caused by high electrode reactivity. Fluorinated solvent-based electrolytes can address this challenge, but they pose environmental and biological hazards. This work reports on the molecular engineering of fluorine (F)-free ethers to mitigate electrode surface reactivity in high-voltage Ni-rich LMBs. By merely extending the alkyl chains of traditional ethers, we effectively reduce the catalytic reactivity of the cathode towards the electrolyte at high voltages, which suppresses the oxidation decomposition of the electrolyte, microstructural defects and rock-salt phase formation in the cathode, and gas release issues. The high-voltage Ni-rich NCM811-Li battery delivers capacity retention of 80 % after 250 cycles with a high Coulombic efficiency of 99.85 %, even superior to that in carbonate electrolytes. Additionally, this strategy facilitates passivation of the Li anode by forming a robust solid-electrolyte interphase, boosting the Li reversibility to 99.11 % with a cycling life of 350 cycles, which outperforms conventional F-free ether electrolytes. Consequently, the lifespan of practical LMBs has been prolonged by over 100 % and 500 % compared to those in conventional carbonate- and ether-based electrolytes, respectively.

Promoter R-Loops Recruit U2AF1 to Modulate Its Phase Separation and RNA Splicing.

R-loops and guanine quadruplexes (G4s) are secondary structures of nucleic acids that are ubiquitously present in cells and are enriched in promoter regions of genes. By employing a bioinformatic approach based on overlap analysis of transcription factor chromatin immunoprecipitation sequencing (ChIP-seq) data sets, we found that many splicing factors, including U2AF1 whose recognition of the 3' splicing site is crucial for pre-mRNA splicing, exhibit pronounced enrichment at endogenous R-loop- and DNA G4-structure loci in promoter regions of human genes. We also revealed that U2AF1 binds directly to R-loops and DNA G4 structures at a low-nM binding affinity. Additionally, we showed the ability of U2AF1 to undergo phase separation, which could be stimulated by binding with R-loops, but not duplex DNA, RNA/DNA hybrid, DNA G4, or single-stranded RNA. We also demonstrated that U2AF1 binds to promoter R-loops in human cells, and this binding competes with U2AF1's interaction with 3' splicing site and leads to augmented distribution of RNA polymerase II (RNAPII) to promoters over gene bodies, thereby modulating cotranscriptional pre-mRNA splicing. Together, we uncovered a group of candidate proteins that can bind to both R-loops and DNA G4s, revealed the direct and strong interactions of U2AF1 with these nucleic acid structures, and established a biochemical rationale for U2AF1's occupancy in gene promoters. We also unveiled that interaction with R-loops promotes U2AF1's phase separation, and our work suggests that U2AF1 modulates pre-mRNA splicing by regulating RNAPII's partition in transcription initiation versus elongation.

Detection of human annexin A1 as the novel N-terminal tag for separation and purification handle.

BACKGROUND: Several fusion tags for separation handle have been developed, but the fused tag for simply and cheaply separating the target protein is still lacking. RESULTS: Separation conditions for the human annexin A1 (hanA1) tagged emerald green fluorescent protein (EmGFP) in Escherichia coli were optimized via precipitation with calcium chloride (CaCl2) and resolubilization with ethylenediamine tetraacetic acid disodium salt (EDTA-Na2). The HanA1-EmGFP absorbing with other three affinity matrix was detected, only it was strongly bound to heparin Sepharose. The separation efficiency of the HanA1-EmGFP was comparable with purification efficiency of the His6-tagged HanA1-EmGFP via metal ion affinity chromatography. Three fluorescent proteins for the EmGFP, mCherry red fluorescent protein and flavin-binding cyan-green fluorescent protein LOV from Chlamydomonas reinhardtii were used for naked-eye detection of the separation and purification processes, and two colored proteins including a red protein for a Vitreoscilla hemoglobin (Vhb), and a brown protein for maize sirohydrochlorin ferrochelatase (mSF) were used for visualizing the separation process. The added EDTA-Na2 disrupted the Fe-S cluster in the mSF, but it showed little impact on heme in Vhb. CONCLUSIONS: The selected five colored proteins were efficient for detecting the applicability of the highly selective hanA1 for fusion separation and purification handle. The fused hanA1 tag will be potentially used for simple and cheap affinity separation of the target proteins in industry and diagnosis.

G3BP1 binds to guanine quadruplexes in mRNAs to modulate their stabilities.

RNA guanine quadruplexes (rG4) assume important roles in post-transcriptional regulations of gene expression, which are often modulated by rG4-binding proteins. Hence, understanding the biological functions of rG4s requires the identification and functional characterizations of rG4-recognition proteins. By employing a bioinformatic approach based on the analysis of overlap between peaks obtained from rG4-seq analysis and those detected in >230 eCLIP-seq datasets for RNA-binding proteins generated from the ENCODE project, we identified a large number of candidate rG4-binding proteins. We showed that one of these proteins, G3BP1, is able to bind directly to rG4 structures with high affinity and selectivity, where the binding entails its C-terminal RGG domain and is further enhanced by its RRM domain. Additionally, our seCLIP-Seq data revealed that pyridostatin, a small-molecule rG4 ligand, could displace G3BP1 from mRNA in cells, with the most pronounced effects being observed for the 3'-untranslated regions (3'-UTR) of mRNAs. Moreover, luciferase reporter assay results showed that G3BP1 positively regulates mRNA stability through its binding with rG4 structures. Together, we identified a number of candidate rG4-binding proteins and validated that G3BP1 can bind directly with rG4 structures and regulate the stabilities of mRNAs.

Kif15 deficiency contributes to depression-like behavior in mice.

Neuropsychiatric disorders have a high incidence worldwide. Kinesins, a family of microtubule-based molecular motor proteins, play essential roles in intracellular and axonal transport. Variants of kinesins have been found to be related to many diseases, including neurodevelopmental/neurodegenerative disorders. Kinesin-12 (also known as Kif15) was previously found to affect the frequency of both directional microtubule transports. However, whether Kif15 deficiency impacts mood in mice is yet to be investigated. In this study, we used the CRISPR/Cas9 method to obtain Kif15-/- mice. In behavioral tests, Kif15-/- female mice exhibited prominent depressive characteristics. Further studies showed that the expression of BDNF was significantly decreased in the frontal cortex, corpus callosum, and hippocampus of Kif15-/- mice, along with the upregulation of Interleukin-6 and Interleukin-1ß in the corpus callosum. In addition, the expression patterns of AnkG were notably changed in the developing brain of Kif15-/- mice. Based on our previous studies, we suggested that this appearance of altered AnkG was due to the maladjustment of the microtubule patterns induced by Kif15 deficiency. The distribution of PSD95 in neurites notably decreased after cultured neurons treated with the Kif15 inhibitor, but total PSD95 protein level was not impacted, which revealed that Kif15 may contribute to PSD95 transportation. This study suggested that Kif15 may serve as a potential target for future depression studies.

The developmental miR-17-92 cluster and the Sfmbt2 miRNA cluster cannot rescue the abnormal embryonic development generated using obstructive epididymal environment-producing sperm in C57BL/6 J mice.

BACKGROUND: Sperm, during epididymal transit, acquires microRNAs(miRNAs), which are crucial for embryonic development. However, whether sperm miRNAs influenced by an obstructive epididymal environment affect embryonic development remains unknown. METHOD: The sham operation and vasectomy were performed in C57BL/6 J mice to create the control group (CON) and the obstructive epididymal environment group(OEE) group, respectively. The morphology of the testis and epididymis was observed using hematoxylin and eosin staining (HE staining) to establish the OEE mice model. The sperm quality test, intracytoplasmic sperm injection (ICSI), and epididymosomes fusion were employed to observe the effect of the obstructive epididymal environment on sperm and resultant embryonic development. The alteration of the sperm small RNA (sRNA) profile was analyzed by sRNA sequencing. RT-qPCR and DNA methylation were applied to observe the effect of obstructive epididymis on the expression of sperm miRNAs. The miRNAs microinjection was used to explore the impacts of sperm miRNAs on embryonic development. RESULTS: We confirmed postoperative 8-week mice as the OEE mice model by examining the morphology of the testis and epididymis. In the OEE group, we observed that sperm quality degraded and the development potential of embryos was reduced, which can be saved by the normal epididymal environment. The sperm sRNA sequencing revealed that the expression of the developmental miR-17-92 cluster and the Sfmbt2 miRNA cluster was downregulated in the OEE group. The expression of these two miRNA clusters in epididymis was also downregulated and regulated by DNA methylation. However, the downregulation of either the miR-17-92 cluster or the Sfmbt2 miRNA cluster in normal zygotes did not impair embryonic development. CONCLUSION: The obstructive epididymal environment influences sperm quality and resultant embryonic development, as well as the abundance of the developmental miR-17-92 cluster and the Sfmbt2 miRNA cluster in sperm, but these miRNA clusters are not the cause of abnormal embryonic development. It implies that epididymis is important in early embryonic development and may play a potential role in sperm epigenome.

Body mass index and pharmacodynamics of target-controlled infusion of propofol: A prospective non-randomized controlled study.

WHAT IS KNOWN AND OBJECTIVE: In our preliminary study, there were large individual variations at sedation levels during propofol target-controlled infusion (TCI). The present study aimed to assess the effects of body mass index (BMI) on the pharmacodynamic index of propofol TCI. METHODS: This prospective, non-randomized controlled trial evaluated 175 female patients undergoing breast lumpectomy. Anesthesia was induced with propofol using the TCI system embedded Schnider model. The effect compartment concentration was set to 3 µg/ml, and the start time of infusion was recorded. When the target concentration reached 3 µg/ml, the patient could not be awakened (Ramsay sedation score ≥4), and when the Bispectral Index (BIS) was <60, the infusion was discontinued, and the time point was recorded. The observation end-point was set at the Observer's Assessment of Alertness/Sedation (OAA/S) score of <4. The correlation between the BMI and the pharmacodynamic index of propofol was evaluated. RESULTS AND DISCUSSION: Propofol induction time was significantly correlated with the BMI (p < 0.001). The induction time of the underweight subjects was 10.14 ± 2.19 min, which was remarkably higher than that of normal weight (6.48 ± 3.44 min) and overweight (4.75 ± 2.53 min) individuals (p < 0.001). There were still significant differences after multivariable-adjusted regressions (p < 0.001). There were no significant differences in recovery time and sedative effect indicators, such as Ramsay score, BIS value, and effect compartment concentration, between the three groups (p > 0.05 for all). WHAT IS NEW AND CONCLUSION: These results suggest that the BMI is one of the critical factors affecting the pharmacodynamic index of propofol TCI, and the induction time decreased progressively with increasing BMI. The Schnider model might underpredict doses of propofol for underweight individuals.

The roles of polymerases ν and θ in replicative bypass of O6- and N2-alkyl-2'-deoxyguanosine lesions in human cells.

Exogenous and endogenous chemicals can react with DNA to produce DNA lesions that may block DNA replication. Not much is known about the roles of polymerase (Pol) ν and Pol θ in translesion synthesis (TLS) in cells. Here we examined the functions of these two polymerases in bypassing major-groove O6-alkyl-2'-deoxyguanosine (O6-alkyl-dG) and minor-groove N2-alkyl-dG lesions in human cells, where the alkyl groups are ethyl, n-butyl (nBu), and, for O6-alkyl-dG, pyridyloxobutyl. We found that Pol ν and Pol θ promote TLS across major-groove O6-alkyl-dG lesions. O6-alkyl-dG lesions mainly induced G→A mutations that were modulated by the two TLS polymerases and the structures of the alkyl groups. Simultaneous ablation of Pol ν and Pol θ resulted in diminished mutation frequencies for all three O6-alkyl-dG lesions. Depletion of Pol ν alone reduced mutations only for O6-nBu-dG, and sole loss of Pol θ attenuated the mutation rates for O6-nBu-dG and O6-pyridyloxobutyl-dG. Replication across the two N2-alkyl-dG lesions was error-free, and Pol ν and Pol θ were dispensable for their replicative bypass. Together, our results provide critical knowledge about the involvement of Pol ν and Pol θ in bypassing alkylated guanine lesions in human cells.

Mitochondrial Transcription Factor A Binds to and Promotes Mutagenic Transcriptional Bypass of O4-Alkylthymidine Lesions.

O2- and O4-alkylated thymidine lesions are known to be poorly repaired and persist in mammalian tissues. To understand how mammalian cells sense the presence and regulate the repair of these lesions, we employed a quantitative proteomic method to discover regioisomeric O2- and O4-n-butylthymidine (O2- and O4-nBudT)-binding proteins. We were able to identify 21 and 74 candidate DNA damage recognition proteins for O2-nBudT- and O4-nBudT-bearing DNA probes, respectively. Among these proteins, DDB1 and DDB2 selectively bind to O2-nBudT-containing DNA, whereas three high-mobility group (HMG) proteins (i.e., HMGB1, HMGB2, and mitochondrial transcription factor A (TFAM)) exhibit preferential binding to O4-nBudT-bearing DNA. We further demonstrated that TFAM binds directly and selectively with O4-alkyldT-harboring DNA, and the binding capacity depends mainly on the HMG box-A domain of TFAM. We also found that TFAM promotes transcriptional mutagenesis of O4-nBudT and O4-pyridyloxobutylthymidine, which is a DNA adduct induced by tobacco-specific N-nitrosamines, in vitro and in human cells. Together, we explored, for the first time, the interaction proteomes of O-alkyldT lesions, and our study expanded the functions of TFAM by revealing its capability in the recognition of O4-alkyldT-bearing DNA and uncovering its modulation of transcriptional mutagenesis of these lesions in human cells.

Berberine impairs coxsackievirus B3-induced myocarditis through the inhibition of virus replication and host pro-inflammatory response.

Berberine (BBR), an isoquinoline alkaloid isolated from Rhizoma coptidis, is reported to possess antiviral activity. Our previous study has shown that BBR alleviates coxsackievirus B3 (CVB3) replication in HeLa cells. However, the anti-CVB3 activity of BBR is still unclear in vivo. In this study, we explored the effect of BBR on CVB3-induced viral myocarditis in mice. These results demonstrated the beneficial effect of BBR on alleviating CVB3-induced myocarditis in vivo, which sheds new light on the utility of BBR as a therapeutic strategy against CVB3-induced viral myocarditis.

Dendrobium huoshanense Polysaccharides Prevent Inflammatory Response of Ulcerative Colitis Rat through Inhibiting the NF-κB Signaling Pathway.

The polysaccharides of the Chinese herbal medicine Dendrobium huoshanense exhibit anti-inflammatory effects in multiple organs through regulating the immune responses. In the present study, we constructed ulcerative colitis (UC) model rats using dextran sulfate sodium to investigate the anti-inflammatory effects of D. huoshanense polysaccharides (DHP). After oral administration of DHP for two weeks, the indices of UC symptoms, including the ratio of colon weight to length, Disease Activity Index (DAI), and Colon Mucosal Damage Index (CMDI), all decreased significantly compared with the UC model group. The histological sections also revealed better cell orders in DHP treatments than in the UC model rats. Moreover, in treatment with high dose of DHP (200 mg/kg), the treatment efficacy arrived the similar levels to those in the treatment with 300 mg/kg sulfasalazine, which is a typical medicine to treat UC. These results indicated that DHP has a high efficacy to treat UC in model rats. Furthermore, serum levels of interleukin-1ß, tumor necrosis factor-α, interleukin-17, and transforming growth factor-ß were assessed using the enzyme linked immunosorbent assay (ELISA) method, and the levels of nuclear factor-κB in colon tissue sections were determined using the immunohistochemical method. The results showed that all these indices decreased significantly after administration of DHP in UC model rats, which might be the mechanisms underlying the DHP-suppressed UC inflammation. Overall, this study indicated that DHP might be directly used to treat UC and is a promising source to develop novel drugs against UC.

Frequent and mild scrotal heat stress in mice epigenetically alters glucose metabolism in the male offspring.

Several studies have reported that health problems occur in assisted reproductive technology (ART)-conceived offspring. Recently, investigations have demonstrated that paternal environmental conditions influence offspring health. However, it is unclear whether the factors that cause male infertility per se affect offspring health and contribute to health problems in ART-born children. Scrotal heat stress represents a common cause for oligoasthenozoospermia, and in these cases, in vitro fertilization-embryo transfer (IVF-ET) is typically recommended for those individuals trying to conceive. We exposed C57BL/6J male mice to frequent and mild scrotal heat stress (fmSHS) (39°C for 30 min once weekly for 5 consecutive wk). Sperm was subjected to IVF-ET with oocytes of untreated C57BL/6J females to produce offspring mice. Glucose intolerance and insulin resistance was observed in the male offspring mice derived from fmSHS-exposed fathers. Islets, after evaluation, remained unchanged. Genes involved in glucose metabolism, especially, those in insulin signaling pathways, showed dysregulation in the liver of the fmSHS-derived male offspring. Differentially methylated regions were found in the sperm of fmSHS-exposed mice by whole genome bisulfite sequencing. Interestingly, abnormal methylation of some genes with altered expression in offspring was observed in both the sperm of fmSHS fathers and the liver of their male offspring. Our results suggest that the factors that cause male infertility can affect male offspring health by an epigenetic mechanism.

Chemical Proteomic Profiling of the Interacting Proteins of Isoprenoid Pyrophosphates.

Isoprenoid pyrophosphates are involved in protein prenylation and assume regulatory roles in cells; however, little is known about the cellular proteins that can interact with isoprenoid pyrophosphates. Here, we devised a chemical proteomic strategy, capitalizing on the use of a desthiobiotin-geranyl pyrophosphate (GPP) acyl phosphate probe for the enrichment and subsequent identification of GPP-binding proteins using liquid chromatography-tandem mass spectrometry (LC-MS/MS). By combining stable isotope labeling by amino acids in cell culture (SILAC) and competitive labeling with low vs high concentrations of GPP probe, with ATP vs GPP acyl phosphate probes, or with the GPP probe in the presence of different concentrations of free GPP, we uncovered a number of candidate GPP-binding proteins. We also discovered, for the first time, histone deacetylase 1 (HDAC1) as a GPP-binding protein. Furthermore, we found that the enzymatic activity of HDAC1 could be modulated by isoprenoid pyrophosphates. Together, we developed a novel chemical proteomic method for the proteome-wide discovery of GPP-binding proteins, which sets the stage for a better understanding about the biological functions of isoprenoids.

Frequent and mild scrotal heat stress impairs embryo development, implantation and offspring sex ratio in mice.

RESEARCH QUESTION: Frequent and mild scrotal heat stress (fmSHS) often occurs in humans. The aim of this study was to determine the impact of fmSHS on natural fertility, the IVF-embryo transfer (IVF-ET) process and the offspring sex ratio. DESIGN: Male mice were randomly divided into four groups: no scrotal heat stress (SHS) (controls) and those subjected to SHS at 37°C, 39°C or 41°C for 30 min once a week for 5 consecutive weeks. The testis, epididymis and sperm quality were assessed to evaluate the effects of different degrees of SHS, to establish an fmSHS model. Then, natural fertility, IVF-embryo transfer (IVF-ET) results, embryo development, offspring sex ratio and the X/Y chromosome-bearing sperm ratio were examined. RESULTS: SHS at 39°C and 41°C caused mild impairment to spermatozoa, leading to a phenotype similar to oligoasthenozoospermia in humans. Given that most SHS conditions in humans are close to body temperature, SHS of 39°C was adopted to build the fmSHS model. fmSHS reduced the fertilization rate, impaired on-time development and reduced the implantation rate of the embryos in the IVF-ET process, but it did not affect the development or function of blastocysts. The fmSHS mice produced more Y chromosome-bearing spermatozoa and propagated more male offspring. CONCLUSIONS: fmSHS not only reduced the fertilization ability of spermatozoa but also influenced their function beyond fertilization, in addition to changing the offspring sex ratio. These results may help to shed new light on the infertility treatment of males with scrotal heat risk and the health concerns of offspring propagated from these fathers.

The effect of body mass index and creatinine clearance on serum trough concentration of vancomycin in adult patients.

BACKGROUND: The aim of this study was to evaluate the influence of patient body mass index (BMI) and estimated creatinine clearance (CrCl) on serum vancomycin concentrations to define a possible optimal dosage regimen in overweight patients based on data obtained during therapeutic drug monitoring. METHODS: This retrospective study used data collected from January 2017 to January 2019. Adult patients (n = 204) received vancomycin treatment at a dose of 1000 mg every 12 h and underwent serum monitoring. Data collected included patient disease category, sex, age, height, weight, vancomycin concentrations, and serum creatinine. The CrCl values were estimated using the Cockcroft-Gault formula. In this study, statistical comparisons were performed on the results of patients according to serum vancomycin concentration. RESULTS: Serum vancomycin concentration was significantly related to BMI (P <  0.001) and CrCl (P <  0.05) in adult patients. Furthermore, the trough serum vancomycin concentration showed a logarithmic correlation with BMI (R = - 0.5108, 95% CI: - 0.6082 to - 0.3982, P <  0.001) and CrCl (R = - 0.5739, 95% CI: - 0.6616 to - 0.4707, P <  0.001). The multivariate analysis showed that BMI and CrCl are independent contributors to the trough vancomycin concentration. Moreover, some of the patients with higher BMI (≥ 24 kg/m2) met the goal trough concentration after an adjustment from 1000 mg every 12 h to 1000 mg every 8 h. CONCLUSIONS: Serum vancomycin concentration decreases progressively with increasing BMI and the augmentation in CrCl in adult patients. The trough concentration of vancomycin should be continuously monitored for patients with a BMI ≥ 24 kg/m2, and the dosage regimen should be adjusted to reach the target trough concentration in these patients to reduce the impact of BMI.

CVB3 Nonstructural 2A Protein Modulates SREBP1a Signaling via the MEK/ERK Pathway.

Coxsackievirus B3 (CVB3) is the predominant pathogen of viral myocarditis. In our previous study, we found that CVB3 caused abnormal lipid accumulation in host cells. However, the underlying mechanisms by which CVB3 disrupts and exploits the host lipid metabolism are not well understood. Sterol regulatory element binding protein 1 (SREBP1) is the major transcriptional factor in lipogenic genes expression. In this study, we demonstrated that CVB3 infection and nonstructural 2A protein upregulated and activated SREBP1a at the transcriptional level. Deletion analysis of SREBP1a promoter revealed that two regions, -1821/-1490 and -312/+217, in this promoter were both required for its activation by 2A. These promoter regions possessed several binding motifs for transcription factor SP1. Next, we used SP1-specific small interfering RNAs (siRNAs) to confirm that SP1 might be the essential factor in SREBP1a upregulation by 2A. Furthermore, we showed that MEK/ERK pathway was involved in the activation of SREBP1a by 2A and that blocking this signaling pathway with the specific inhibitor U0126 attenuated SREBP1a activation and lipid accumulation by 2A. Finally, we showed that inhibition of SREBP1 with siRNAs attenuated lipid accumulation induced by CVB3 infection and reduced virus replication. Moreover, inhibition of the MEK/ERK pathway also led to reduction of SREBP1a activation, lipid accumulation, and virus replication during CVB3 infection. Taken together, these data demonstrate that CVB3 nonstructural 2A protein activates SREBP1a at the transcription level through a mechanism involving MEK/ERK signaling pathway and SP1 transcription factor, which promotes cellular lipid accumulation and benefits virus replication.IMPORTANCE Coxsackievirus B3 (CVB3) infection is the leading cause of viral myocarditis, but effective vaccines and antiviral therapies against CVB3 infection are still lacking. It is important to understand the precise interactions between host and virus for the rational design of effective therapies. During infection, CVB3 disrupts and exploits host lipid metabolism to promote excessive lipid accumulation, which benefits virus replication. SREBP1 is the master regulator of cellular lipid metabolism. Here, we report that one of the viral nonstructural proteins, 2A, upregulates and activates SREBP1a. Furthermore, we find that inhibition of SREBP1 decreases CVB3 virus replication. These results reveal the regulation of SREBP1a expression by 2A and the roles of SREBP1 in lipid accumulation and viral replication during CVB3 infection. Our findings provide a new insight into CVB3 host interactions and inform a potential novel therapeutic target for this important pathogen.

Facile liquid-phase deposition synthesis of titania-coated magnetic sporopollenin for the selective capture of phosphopeptides.

Titania-grafted magnetic sporopollenin is synthesized by the liquid-phase deposition (LPD) technique, characterized by SEM, EDX, and nitrogen adsorption porosimetry, and used for the selective enrichment of phosphorylated peptides. The material is low cost because of easier availability of pollens and has rich surface chemistry which enables strong attachment of titania onto magnetic sporopollenin. The material shows higher selectivity of 1:1000 with ß-casein spiked in BSA. Higher sensitivity of 10 fmol is recorded for phosphopeptides from standard ß-casein digest. Twenty phosphorylated peptides are enriched from milk digest and four endogenous phosphopeptides from diluted human serum. The magnetic property of titania-coated magnetic sporopollenin facilitates the fast and effective isolation of phosphopeptides from complex mixtures through external magnet. The material is finally applied to tryptic digest of rat brain cell lysate for phosphopeptide enrichment where 2718 phosphopeptides are identified by using LC-MS/MS with C18 column. Titania-coated magnetic sporopollenin captures both mono-phosphorylated (2489) and multi-phosphorylated peptides (229) due to strong affinity of TiO2 with phosphates. TiO2-coated magnetic material also shows better enrichment efficiency in comparison to commercial TiO2. Graphical abstract.