Substitutions in the nonactive site of the passenger domain on the activity of Haemophilus influenzae immunoglobulin A1 protease.

Immunoglobulin A1 (IgA1) protease is a critical virulence factor of Haemophilus influenzae that facilitates bacterial mucosal infection. This study investigates the effect of iga gene polymorphism on the enzymatic activity of H. influenzae IgA1 protease. The IgA1 protease activity was examined in the H. influenzae Rd KW20 strain and 51 isolates. Genetic variations in iga and deduced amino acid substitutions affecting IgA1 protease activity were assessed. Machine learning tools and functional complementation assays were used to analyze the effects of identified substitutions on the stability and activity of IgA1 protease, respectively. All 51 isolates exhibited similar iga expression levels. No igaB expression was detected. According to comparisons with the reference Rd KW20 strain, four substitutions in the protease domain, 26 in the nonprotease passenger domain, and two in the ß-barrel domain were associated with the change in IgA1 protease activity. No substitutions in the catalytic site of IgA1 protease were observed. Logistic regression, receiver operating characteristic curves, Venn diagrams, and protein stability analyses revealed that the substitutions Asn352Lys, Pro353Ala, Lys356Asn, Gln916Lys, and Gly917Ser, which were located in the nonactive site of the passenger domain, were associated with decreases in IgA1 protease activity and stability, whereas Asn914Lys was associated with an increase in these events. Functional complementation assays revealed that the Asn914Lys substitution increased IgA1 protease activity in the Rd KW20 strain. This study identified substitutions in the nonactive site of the passenger domain that affect both the activity and stability of H. influenzae IgA1 protease.

Membranome-based identification of amino acid substitution in Haemophilus influenzae multidrug efflux pump HmrM for reduced chloramphenicol susceptibility.

A decreased chloramphenicol susceptibility in Haemophilus influenzae is commonly caused by the activity of chloramphenicol acetyltransferases (CATs). However, the involvement of membrane proteins in chloramphenicol susceptibility in H. influenzae remains unclear. In this study, chloramphenicol susceptibility testing, whole-genome sequencing, and analyses of membrane-related genes were performed in 51 H. influenzae isolates. Functional complementation assays and structure-based protein analyses were conducted to assess the effect of proteins with sequence substitutions on the minimum inhibitory concentration (MIC) of chloramphenicol in CAT-negative H. influenzae isolates. Six isolates were resistant to chloramphenicol and positive for type A-2 CATs. Of these isolates, A3256 had a similar level of CAT activity but a higher chloramphenicol MIC relative to the other resistant isolates; it also had 163 specific variations in 58 membrane genes. Regarding the CAT-negative isolates, logistic regression and receiver operator characteristic curve analyses revealed that 48T > G (Asn16Lys), 85 C > T (Leu29Phe), and 88 C > A (Leu30Ile) in HI_0898 (emrA), and 86T > G (Phe29Cys) and 141T > A (Ser47Arg) in HI_1177 (artM) were associated with enhanced chloramphenicol susceptibility, whereas 997G > A (Val333Ile) in HI_1612 (hmrM) was associated with reduced chloramphenicol susceptibility. Furthermore, the chloramphenicol MIC was lower in the CAT-negative isolates with EmrA-Leu29Phe/Leu30Ile or ArtM-Ser47Arg substitution and higher in those with HmrM-Val333Ile substitution, relative to their counterparts. The Val333Ile substitution was associated with enhanced HmrM protein stability and flexibility and increased chloramphenicol MICs in CAT-negative H. influenzae isolates. In conclusion, the substitution in H. influenzae multidrug efflux pump HmrM associated with reduced chloramphenicol susceptibility was characterised.

Cell-Penetrating Peptides for Use in Development of Transgenic Plants.

Genetically modified plants and crops can contribute to remarkable increase in global food supply, with improved yield and resistance to plant diseases or insect pests. The development of biotechnology introducing exogenous nucleic acids in transgenic plants is important for plant health management. Different genetic engineering methods for DNA delivery, such as biolistic methods, Agrobacterium tumefaciens-mediated transformation, and other physicochemical methods have been developed to improve translocation across the plasma membrane and cell wall in plants. Recently, the peptide-based gene delivery system, mediated by cell-penetrating peptides (CPPs), has been regarded as a promising non-viral tool for efficient and stable gene transfection into both animal and plant cells. CPPs are short peptides with diverse sequences and functionalities, capable of agitating plasma membrane and entering cells. Here, we highlight recent research and ideas on diverse types of CPPs, which have been applied in DNA delivery in plants. Various basic, amphipathic, cyclic, and branched CPPs were designed, and modifications of functional groups were performed to enhance DNA interaction and stabilization in transgenesis. CPPs were able to carry cargoes in either a covalent or noncovalent manner and to internalize CPP/cargo complexes into cells by either direct membrane translocation or endocytosis. Importantly, subcellular targets of CPP-mediated nucleic acid delivery were reviewed. CPPs offer transfection strategies and influence transgene expression at subcellular localizations, such as in plastids, mitochondria, and the nucleus. In summary, the technology of CPP-mediated gene delivery provides a potent and useful tool to genetically modified plants and crops of the future.

Curcumin analog HO-3867 triggers apoptotic pathways through activating JNK1/2 signalling in human oral squamous cell carcinoma cells.

Human oral squamous cell carcinoma (OSCC) is the common head and neck malignancy in the world. While surgery, radiotherapy and chemotherapy are emerging as the standard treatment for OSCC patients, the outcome is limited to the recurrence and side effects. Therefore, patients with OSCC require alternative strategies for treatment. In this study, we aimed to explore the therapeutic effect and the mode of action of the novel curcumin analog, HO-3867, against human OSCC cells. We analysed the cytotoxicity of HO-3867 using MTT assay. In vitro mechanic studies were performed to determine whether MAPK pathway is involved in HO-3867 induced cell apoptosis. As the results, we found HO-3867 suppressed OSCC cells growth effectively. The flow cytometry data indicate that HO-3867 induce the sub-G1 phase. Moreover, we found that HO-3867 induced cell apoptosis by triggering formation of activated caspase 3, caspase 8, caspase 9 and PARP. After dissecting MAPK pathway, we found HO-3867 induced cell apoptosis via the c-Jun N-terminal kinase (JNK)1/2 pathway. Our results suggest that HO-3867 is an effective anticancer agent as its induction of cell apoptosis through JNK1/2 pathway in human oral cancer cells.

TIMP3 Gene Polymorphisms of -1296 T > C and -915 A > G Increase the Susceptibility to Arsenic-Induced Skin Cancer: A Cohort Study and In Silico Analysis of Mutation Impacts

Long-term exposure to arsenic may induce several human cancers, including non-melanoma skin cancer. The tissue inhibitor of metalloproteinase (TIMP)-3, encoded by the TIMP3 gene, may inhibit tumor growth, invasion, and metastasis of several cancer types. In this study, we aimed to investigate effects of the TIMP3 -1296 T > C (rs9619311) and -915 A > G (rs2234921) single-nucleotide polymorphisms (SNPs) on skin cancer risk in an arsenic-exposed population, and to evaluate the influence of allele-specific changes by an in silico analysis. In total, 1078 study participants were followed up for a median of 15 years for newly diagnosed skin cancer. New cases were identified through linkage to the National Cancer Registry of Taiwan. A Cox regression analysis was used to evaluate the effects of TIMP3 variants. Transcription factor (TF) profiling of binding sites of allele-specific changes in SNPs was conducted using the JASPAR scan tool. We observed borderline associations between TIMP3 genotypes and skin cancer risk. However, when combined with high arsenic exposure levels, the rs9619311 C allele, rs2234921 G allele, or C-G haplotype groups exhibited a greater risk of developing skin cancer compared to the respective common homozygous genotype group. The in silico analysis revealed several TF motifs located at or flanking the two SNP sites. We validated that the C allele of rs9619311 attenuated the binding affinity of BACH2, MEIS2, NFE2L2, and PBX2 to the TIMP3 promoter, and that the G allele of rs2234921 reduced the affinity of E2F8 and RUNX1 to bind to the promoter. Our findings suggest significant modifications of the effect of the association between arsenic exposure and skin cancer risk by the TIMP3 rs9619311 and rs2234921 variants. The predicted TFs and their differential binding affinities to the TIMP3 promoter provide insights into how TIMP3 interacts with arsenic through TFs in skin cancer formation.

Low-Voltage Electricity-Associated Burn Damage of Lung Parenchyma: Case Report and Literature Review.

BACKGROUND: Electrical injuries are common in daily life. The severity of electrical injury depends on the electric current, and assessing electrical damage is difficult because there appears to be no correlation between skin burns and visceral injury. We report a case of bilateral lung injury with pulmonary hemorrhage after exposure to low-voltage electricity. CASE REPORT: A 23-year-old man was shocked by a low-voltage (110 V) electric current while at work. He had temporary loss of consciousness and twitching in the extremities, but soon regained consciousness and spontaneously stopped twitching. Electrical burn wounds were discovered on his back and forehead. Dyspnea and hemoptysis were noted. A computed tomography scan of the chest revealed patchy infiltration and consolidation of both lungs. The patient received treatment of tranexamic acid and prophylactic antibiotics for electricity-induced lung injury and pulmonary hemorrhage. Resolution of chest radiograph abnormalities was recorded on day 7. The mild dyspnea ceased approximately 2 weeks later. WHY SHOULD AN EMERGENCY PHYSICIAN BE AWARE OF THIS?: Electricity-induced lung injury should be considered in patients with electrical injury through a suspicious electrical current transmission pathway, respiratory symptoms, and corresponding imaging findings. Pulmonary complications can be serious and require early intervention.

EAT-Rice: A predictive model for flanking gene expression of T-DNA insertion activation-tagged rice mutants by machine learning approaches.

T-DNA activation-tagging technology is widely used to study rice gene functions. When T-DNA inserts into genome, the flanking gene expression may be altered using CaMV 35S enhancer, but the affected genes still need to be validated by biological experiment. We have developed the EAT-Rice platform to predict the flanking gene expression of T-DNA insertion site in rice mutants. The three kinds of DNA sequences including UPS1K, DISTANCE, and MIDDLE were retrieved to encode and build a forecast model of two-layer machine learning. In the first-layer models, the features nucleotide context (N-gram), cis-regulatory elements (Motif), nucleotide physicochemical properties (NPC), and CG-island (CGI) were used to build SVM models by analysing the concealed information embedded within the three kinds of sequences. Logistic regression was used to estimate the probability of gene activation which as feature-encoding weighting within first-layer model. In the second-layer models, the NaiveBayesUpdateable algorithm was used to integrate these first layer-models, and the system performance was 88.33% on 5-fold cross-validation, and 79.17% on independent-testing finally. In the three kinds of sequences, the model constructed by Middle had the best contribution to the system for identifying the activated genes. The EAT-Rice system provided better performance and gene expression prediction at further distances when compared to the TRIM database. An online server based on EAT-rice is available at http://predictor.nchu.edu.tw/EAT-Rice.

GasPhos: Protein Phosphorylation Site Prediction Using a New Feature Selection Approach with a GA-Aided Ant Colony System.

Protein phosphorylation is one of the most important post-translational modifications, and many biological processes are related to phosphorylation, such as DNA repair, transcriptional regulation and signal transduction and, therefore, abnormal regulation of phosphorylation usually causes diseases. If we can accurately predict human phosphorylation sites, this could help to solve human diseases. Therefore, we developed a kinase-specific phosphorylation prediction system, GasPhos, and proposed a new feature selection approach, called Gas, based on the ant colony system and a genetic algorithm and used performance evaluation strategies focused on different kinases to choose the best learning model. Gas uses the mean decrease Gini index (MDGI) as a heuristic value for path selection and adopts binary transformation strategies and new state transition rules. GasPhos can predict phosphorylation sites for six kinases and showed better performance than other phosphorylation prediction tools. The disease-related phosphorylated proteins that were predicted with GasPhos are also discussed. Finally, Gas can be applied to other issues that require feature selection, which could help to improve prediction performance. GasPhos is available at http://predictor.nchu.edu.tw/GasPhos.

KStable: A Computational Method for Predicting Protein Thermal Stability Changes by K-Star with Regular-mRMR Feature Selection.

Thermostability is a protein property that impacts many types of studies, including protein activity enhancement, protein structure determination, and drug development. However, most computational tools designed to predict protein thermostability require tertiary structure data as input. The few tools that are dependent only on the primary structure of a protein to predict its thermostability have one or more of the following problems: a slow execution speed, an inability to make large-scale mutation predictions, and the absence of temperature and pH as input parameters. Therefore, we developed a computational tool, named KStable, that is sequence-based, computationally rapid, and includes temperature and pH values to predict changes in the thermostability of a protein upon the introduction of a mutation at a single site. KStable was trained using basis features and minimal redundancy-maximal relevance (mRMR) features, and 58 classifiers were subsequently tested. To find the representative features, a regular-mRMR method was developed. When KStable was evaluated with an independent test set, it achieved an accuracy of 0.708.

Urgent Management of Penetrating Ocular Injury: A Case Report and Review of the Literature.

Ocular globe injury is a severe ophthalmic emergency that requires immediate attention in the emergency department. In this case report, we present a 35-year-old male who suffered a penetrating ocular injury and globe rupture caused by a nail puncture. The patient presented with severe pain and visual loss and was treated with tetanus vaccination, empirical antibiotics, and pain control, followed by an urgent orbital computed tomography (CT) scan and consultation with an ophthalmologist. The CT scan revealed a retained nail in the ocular space, and an urgent operation was performed to repair the eyeball rupture, remove the intraocular foreign body, and perform an anterior vitrectomy. The patient was discharged 6 days after the operation with a visual acuity of 20/400 and an ocular trauma score of 34. This case highlights the importance of initial emergency physician decision-making and the need for a thorough history-taking and examination when encountering penetrating ocular injuries.

Using ensemble learning and hierarchical strategy to predict the outcomes of ESWL for upper ureteral stone treatment.

Urinary tract stones are a common and frequently recurring medical issue. Accurately predicting the success rate after surgery can help avoid ineffective medical procedures and reduce unnecessary healthcare costs. This study collected data from patients with upper ureter stones who underwent extracorporeal shock wave lithotripsy, including cases of successful as well as unsuccessful stone removal after the first and second lithotripsy procedures, and constructed prediction systems for the outcomes of the first and second lithotripsy procedures. Features were extracted from three categories of information: patient characteristics, stone characteristics, and extracorporeal shock wave lithotripsy machine data, and additional features were created using Feature Creation. Finally, the impact of features on the models was analyzed using six methods to calculate feature importance. Our prediction model for the first lithotripsy, selected from among 43 methods and seven ensemble learning techniques, achieves an AUC of 0.91. For the second lithotripsy, the AUC reaches 0.76. The results indicate that the detailed and binary information provided by patients regarding their history of stone experiences contributes differently to the predictive accuracy of the first and second lithotripsy procedures. The prediction tool is available at https://predictor.isu.edu.tw/ks.

Therapeutic Options Targeting the Ataxia-Telangiectasia Mutated (ATM)-mediated DNA Damage Response, Macropinocytosis, and Adaptive Immunity in Ovarian Cancer.

Ataxia-telangiectasia mutated (ATM) is a pivotal protein with versatile kinase activity that responds to DNA damage. While its well-established role as a DNA repair protein is widely recognized, the understanding of its noncanonical functions in ovarian cancer remains limited. Numerous studies have investigated the potential of targeting ATM for ovarian cancer treatment. In addition to its involvement in homologous recombination repair (HRR), an increasing body of research suggests that ATM plays a role in cellular metabolism and adaptive immunity. This review focuses on the current evidence and provides a perspective on how targeting ATM in ovarian cancer can address HRR-deficient genotypes, influence macropinocytosis, and enhance immune checkpoint blockade (ICB) therapy. It underscores the diverse avenues through which targeting ATM is a potential tailored treatment for ovarian cancer.

iStable: off-the-shelf predictor integration for predicting protein stability changes.

BACKGROUND: Mutation of a single amino acid residue can cause changes in a protein, which could then lead to a loss of protein function. Predicting the protein stability changes can provide several possible candidates for the novel protein designing. Although many prediction tools are available, the conflicting prediction results from different tools could cause confusion to users. RESULTS: We proposed an integrated predictor, iStable, with grid computing architecture constructed by using sequence information and prediction results from different element predictors. In the learning model, several machine learning methods were evaluated and adopted the support vector machine as an integrator, while not just choosing the majority answer given by element predictors. Furthermore, the role of the sequence information played was analyzed in our model, and an 11-window size was determined. On the other hand, iStable is available with two different input types: structural and sequential. After training and cross-validation, iStable has better performance than all of the element predictors on several datasets. Under different classifications and conditions for validation, this study has also shown better overall performance in different types of secondary structures, relative solvent accessibility circumstances, protein memberships in different superfamilies, and experimental conditions. CONCLUSIONS: The trained and validated version of iStable provides an accurate approach for prediction of protein stability changes. iStable is freely available online at: http://predictor.nchu.edu.tw/iStable.

ATM inhibition drives metabolic adaptation via induction of macropinocytosis.

Macropinocytosis is a nonspecific endocytic process that may enhance cancer cell survival under nutrient-poor conditions. Ataxia-Telangiectasia mutated (ATM) is a tumor suppressor that has been previously shown to play a role in cellular metabolic reprogramming. We report that the suppression of ATM increases macropinocytosis to promote cancer cell survival in nutrient-poor conditions. Combined inhibition of ATM and macropinocytosis suppressed proliferation and induced cell death both in vitro and in vivo. Supplementation of ATM-inhibited cells with amino acids, branched-chain amino acids (BCAAs) in particular, abrogated macropinocytosis. Analysis of ATM-inhibited cells in vitro demonstrated increased BCAA uptake, and metabolomics of ascites and interstitial fluid from tumors indicated decreased BCAAs in the microenvironment of ATM-inhibited tumors. These data reveal a novel basis of ATM-mediated tumor suppression whereby loss of ATM stimulates protumorigenic uptake of nutrients in part via macropinocytosis to promote cancer cell survival and reveal a potential metabolic vulnerability of ATM-inhibited cells.

Mitochondrial ROS Triggers KIN Pathogenesis in FAN1-Deficient Kidneys.

Karyomegalic interstitial nephritis (KIN) is a genetic adult-onset chronic kidney disease (CKD) characterized by genomic instability and mitotic abnormalities in the tubular epithelial cells. KIN is caused by recessive mutations in the FAN1 DNA repair enzyme. However, the endogenous source of DNA damage in FAN1/KIN kidneys has not been identified. Here we show, using FAN1-deficient human renal tubular epithelial cells (hRTECs) and FAN1-null mice as a model of KIN, that FAN1 kidney pathophysiology is triggered by hypersensitivity to endogenous reactive oxygen species (ROS), which cause chronic oxidative and double-strand DNA damage in the kidney tubular epithelial cells, accompanied by an intrinsic failure to repair DNA damage. Furthermore, persistent oxidative stress in FAN1-deficient RTECs and FAN1 kidneys caused mitochondrial deficiencies in oxidative phosphorylation and fatty acid oxidation. The administration of subclinical, low-dose cisplatin increased oxidative stress and aggravated mitochondrial dysfunction in FAN1-deficient kidneys, thereby exacerbating KIN pathophysiology. In contrast, treatment of FAN1 mice with a mitochondria-targeted ROS scavenger, JP4-039, attenuated oxidative stress and accumulation of DNA damage, mitigated tubular injury, and preserved kidney function in cisplatin-treated FAN1-null mice, demonstrating that endogenous oxygen stress is an important source of DNA damage in FAN1-deficient kidneys and a driver of KIN pathogenesis. Our findings indicate that therapeutic modulation of kidney oxidative stress may be a promising avenue to mitigate FAN1/KIN kidney pathophysiology and disease progression in patients.

Characterization of the temporary anion states on perfluoroalkanes via stabilized Koopmans' theorem in long-range corrected density functional theory.

The stabilized Koopmans' theorem (SKT) in long-range corrected density functional theory is used to characterize the temporary anion states of perfluoro-n-alkanes (n-PFAs) from C(2) to C(5), and perfluorocycloalkanes (c-PFAs) from C(3) to C(4). In this approach, stabilization is accomplished by varying the exponents of appropriate diffuse functions. The energies of temporary anion states are then identified by investigating the relationship between the resultant eigenvalues and scale parameter. The characteristics of resonance orbitals are also examined. For the lowest unfilled orbitals of perfluoroalkanes, results indicate that they are mainly from the π-bonding interactions between all neighboring C atoms. In addition, their energies decrease as the sizes of the perfluoroalkanes increase. Moreover, the energies of the c-C(3)F(6)/c-C(4)F(8) are lower than those of the corresponding n-C(3)F(8)/n-C(4)F(10). When compared with experimental data, our SKT calculations can yield conformable results. Thus, this SKT approach can provide more information on the resonance states of perfluoroalkanes.

Energies and lifetimes of temporary anion states of chloromethanes by stabilized Koopmans' theorem in long-range corrected density functional theory.

To investigate the resonance energies and lifetimes of temporary anion states of chloromethanes, long-range corrected density functional theory is adopted in this article. Their values are determined by calculating the density of resonance states via the stabilized Koopmans' theorem. The characteristics of these resonance orbitals are also analyzed. By comparing with experimental values and previous theoretical calculations, our method can yield not only conformable results but also more complete information on the resonance states.