The variations in gene expression of GAPDH in Ocimum basilicum cultivars under drought-induced stress conditions.

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) holds a pivotal role within the glycolytic pathway of higher plants. It has garnered attention as a significant target protein in instances of oxidative stress, where it can engage in thiolation reactions within its active site. Numerous genes encoding cytosolic iterations of GAPDH have been identified and analyzed in specific plant species. This investigation was conducted to gain insights into GAPDH's function amidst drought-induced stress. Within this framework, the basil plant (Ocimum basilicum) was chosen for focused exploration, encompassing the cloning of the comprehensive cDNA of basil GAPDH (ObGAPDH) and scrutinizing its patterns of expression. The complete sequence of Ob-GAPDH spanned 1315 base pairs. The resultant protein derived from this sequence comprised 399 amino acids, projecting a molecular weight of approximately 42.54 kDa and an isoelectric point (pI) of 6.01. An examination of the evolutionary connections among various GAPDH proteins unveiled ObGAPDH's shared lineage with GAPDH proteins sourced from other plants, such as Salvia splendens and Sesamum indicum. Furthermore, computational methodologies were harnessed to predict the potential oxidative role of ObGAPDH in response to external signals. Molecular docking simulations illuminated the interaction between ObGAPDH and hydrogen peroxide (H2O2) as a ligand. Scrutinizing the expression patterns of the ObGAPDH gene under conditions of water scarcity stress brought to light diverse levels of transcriptional activity. Collectively, these findings underscore the notion that the regulation of ObGAPDH expression is contingent upon both the specific plant cultivar and the presence of stress stemming from drought conditions.

The fluctuations of expression profiles of critical genes in the miRNA maturation process and pro-and anti-inflammatory cytokines in the pathogenesis and progression of multiple sclerosis.

BACKGROUND: Multiple sclerosis (MS) is a central nervous system disease known for immune-mediated demyelination, inflammatory, and neurodegeneration symptoms. Discovering molecular biomarkers to classify RRMS and SPMS patients, monitor the disease activity, and response to particular treatments is one area that has received notable attraction. MicroRNA (miRNA), a single-stranded non-coding RNA molecule, is a significant regulator of gene expression recruited in pathogenic mechanisms in diverse diseases, especially cancer and MS. Also, the relapsing-remitting features of MS exhibit that both inflammatory and anti-inflammatory cytokines are effective in the progression of the disease over time. METHODS AND RESULTS: It was assessed the expression patterns of the genes (Drosha, Pasha (DGCR8), and Dicer ) encoding the critical enzymes in the processing steps of miRNA maturation and major pro-inflammatory and anti-inflammatory cytokines (IFN-α, IFN-ß, and IL-6) in blood cells of 40 MS patients (two groups of 10 men and women in both clinical courses of RR and SPMS patients) in comparison with 20 healthy control group (10 males and 10 females). The highest transcription activity of Drosha was observed for RRMS patients (4.2 and 3.6-fold, respectively), and the expression ratio was down regulated in male and female patients with SPMS (3.9- and 3.1-fold, respectively). Considering the studied cytokines, the increase in expression ratio of IL-6 in SPMS patients and the decrease in transcript abundance of INF-α, and INF-ß cytokines are consistent with the progression of the disease. CONCLUSIONS: Our findings showed that the high and low transcriptional levels of the considered genes seem to be effective in the pathogenesis and progression of MS.

Genome-wide identification and expression profile analysis of metal tolerance protein gene family in Eucalyptus grandis under metal stresses.

Metal tolerance proteins (MTPs) as Me2+/H+(K+) antiporters participate in the transport of divalent cations, leading to heavy metal stress resistance and mineral utilization in plants. In the present study, to obtain better knowledge of the biological functions of the MTPs family, 20 potential EgMTPs genes were identified in Eucalyptus grandis and classified into seven groups belonging to three cation diffusion facilitator groups (Mn-CDFs, Zn/Fe-CDFs, and Zn-CDFs) and seven groups. EgMTP-encoded amino acids ranged from 315 to 884, and most of them contained 4-6 recognized transmembrane domains and were clearly prognosticated to localize into the cell vacuole. Almost all EgMTP genes experienced gene duplication events, in which some might be uniformly distributed in the genome. The numbers of cation efflux and the zinc transporter dimerization domain were highest in EgMTP proteins. The promoter regions of EgMTP genes have different cis-regulatory elements, indicating that the transcription rate of EgMTP genes can be a controlled response to different stimuli in multiple pathways. Our findings provide accurate perception on the role of the predicted miRNAs and the presence of SSR marker in the Eucalyptus genome and clarify their functions in metal tolerance regulation and marker-assisted selection, respectively. Gene expression profiling based on previous RNA-seq data indicates a probable function for EgMTP genes during development and responses to biotic stress. Additionally, the upregulation of EgMTP6, EgMTP5, and EgMTP11.1 to excess Cd2+ and Cu2+ exposure might be responsible for metal translocation from roots to leaves.

In vitro analysis of green synthesized CuO nanoparticles using Tanacetum parthenium extract for multifunctional applications.

Tanacetum parthenium L. is a popular traditional medicinal plant that the role of presence of particular phytochemical compounds are still unconsidered particularly in the bio-nano researches. Here, for the first time, the green fabrication of CuO NPs using Tanacetum parthenium L. extract was performed and assessed for the antimicrobial, cytotoxicity, and dye degradation activities. Characterization of CuO NPs was done by UV-visible spectra, XRD, FT-IR, TEM, and EDX. The synthesized CuO NPs possess a crystalline nature, a functional group that resembles T. parthenium, with a spherical shape particle with an average size of 28 nm. EDX confirmed CuO NPs formation. The CuO NPs showed excellent antimicrobial activity against tested microorganisms. The cytotoxicity of CuO NPs was demonstrated the concentration-dependent inhibition of the growth against both cancer and normal cell lines. The results exhibited concentration-dependent inhibition of the growth of Hela, A 549, and MCF7 cancer cells (IC50 = 65.0, 57.4, and 71.8 µg/mL, respectively), which were statistically significant comparing control cells (IC50 = 226.1 µg/mL). Furthermore, we observed that CuO NPs-induced programmed cell death in the cancer cells were mediated with the downregulation of Bcl2 and upregulation of bax, caspase-3. CuO NPs were verified to be a superb catalyst as they had excellent activity for the degradation of 99.6%, 98.7%, 96.6%, and 96.6% of Congo red, methylene blue, methylene orange, and rhodamine B as industrial dyes in 3, 6.5, 6.5, and 6.5 min, respectively. Overall, the present study nominates T. parthenium as a proper bio-agent in the biosynthesis of CuO NPs with powerful catalytic and antimicrobial activities as well as a cancer treatment.

Comparison and monitoring of antibody response in convalescent and healthy vaccinated individuals against RBD and PCS of SARS-CoV-2 spike protein.

The prevalence of SARS-CoV-2 as a global health threat has called for population-wide vaccination to curb COVID-19. Hence, the World Health Organization (WHO) has approved several platforms of SARS-CoV-2 vaccines for emergency use. Therefore, a more comprehensive study on the immune response induced by vaccines in diverse individuals is still required. Here, we expressed a local variant of SARS-CoV-2 receptor-binding domain (RBD) and protease cleavage site (PCS), playing a vital role in binding and fusion in Rosetta (DE3). We then characterized it through SDS-PAGE analysis and western blotting. Moreover, we compared and monitored ChAdOx1 nCoV-19 vaccination-induced antibody response in convalescent and healthy vaccinated individuals after the first and second vaccine doses through serologic assay against RBD and PCS, which have not yet been compared. We investigated a cohort of 100 sera samples; based on our parameters, 25 serum samples were selected as convalescent samples and 25 serum samples as healthy samples for comparison. These findings demonstrate that most of the convalescent sera show more reactivity with PCS (80%) than with RBD (56%). Interestingly, IgG antibody response against PCS was more significant in both pre- and post-vaccination in convalescent individuals than in healthy individuals. Indeed, anti-RBD antibody titers were most significant in pre-vaccination and post-first vaccination in convalescent individuals than in healthy individuals and not in pre-vaccination and post-second vaccination. Besides monitoring IgG antibody response against COVID-19, these findings could shed light on the progress, assessment, and efficacy evaluation of SARS-CoV-2 vaccines.Communicated by Ramaswamy H. Sarma.

In silico analysis of the conserved surface-exposed epitopes to design novel multiepitope peptide vaccine for all variants of the SARS-CoV-2.

Recently the prevalence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a pervasive threat to generic health. The SARS-CoV-2 spike (S) glycoprotein plays a fundamental role in binds and fusion to the angiotensin-converting enzyme 2 (ACE2). The multi-epitope peptide vaccines would be able to elicit both long-lasting humoral and cellular immune responses, resulting the eliminating SARS-CoV-2 infections as asymptomatic patients are in large numbers. Recently, the omicron variant of the SARS-CoV-2 became a variant of concern that contained just 15-point mutations in the receptor-binding domain of the spike protein. In order to eliminate new evidence on coronavirus variants of concern detected through epidemic intelligence, the conserved epitopes of the receptor-binding domain (RBD) and spike cleavage site is the most probable target for vaccine development to inducing binds and fusion inhibitors neutralizing antibodies respectively. In this study, we utilized bioinformatics tools for identifying and analyzing the spike (S) glycoprotein sequence, e.g. the prediction of the potential linear B-cell epitopes, B-cell multi­epitope design, secondary and tertiary structures, physicochemical properties, solubility, antigenicity, allergenicity, the molecular docking and molecular dynamics simulation for the promising vaccine candidate against all variant of concern of SARS-CoV-2. Among the epitopes of the RBD region are surface-exposed epitopes SVYAWNRKRISNCV and ATRFASVYAWNRKR as the conserved sequences in all variants of concern can be a good candidate to induce an immune response.Communicated by Ramaswamy H. Sarma.

An Insight Into Detection Pathways/Biosensors of Highly Infectious Coronaviruses.

The outbreak of COVID-19 pandemic and its consequences have inflicted a substantial damage on the world. In this study, it was attempted to review the recent coronaviruses appeared among the human being and their epidemic/pandemic spread throughout the world. Currently, there is an inevitable need for the establishment of a quick and easily available biosensor for tracing COVID-19 in all countries. It has been known that the incubation time of COVID-19 lasts about 14 days and 25% of the infected individuals are asymptomatic. To improve the ability to determine SARS-CoV-2 precisely and reduce the risk of eliciting false-negative results produced by mutating nature of coronaviruses, many researchers have established a real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay using mismatch-tolerant molecular beacons as multiplex real-time RT-PCR to distinguish between pathogenic and non-pathogenic strains of coronaviruses. The possible mechanisms and pathways for the detection of coronaviruses by biosensors have been reviewed in this study.

Water deficit stress responses of monoterpenes and sesquiterpenes in different Iranian cultivars of basil.

Ocimum basilicum, a popular aromatic plant, contains aromatic terpenes of terpenoids with in vivo and in vitro verified cytotoxicity. Considering the characteristics and potential of its utilization, it would be attractive to reveal its regulation and biosynthesis, originally at the molecular level under water deficit stress. For this aim, for the first time, the gene encoding the enzyme involved in the end step of the MEP biosynthetic pathways (HDR) was cloned, and the accumulation ratio of linalool, germacrene D and γ-cadinene compounds as well as the expression trait of four critical genes (i.e., HDR, LinS, GerS, and GadS) was assessed under water deficit stress in three Iranian cultivars of basil. The highest value of linalool and γ-cadinene were detected for Cultivar 1 (Cult. 1) under mild stress (W1; 52.6 and 21.1%), while insignificant amounts were obtained for Cultivar 3 (Cult. 3). The germacrene D level of Cultivar 2 (Cult. 2) increased under severe and moderate water stresses as compared with mild water deficit stress. Apart from some expectation, all the studied genes demonstrated divergent transcription ratios under water deficit stress. Principal component analyses (PCA) showed that the relative water content (RWC) and HDR gene expression correlated significantly with essential oil components and gene expression in Cult. 1 and 2, which could represent an elevated demand for corresponding metabolites in the plant tissues. The present work elaborates on the regulation of the mentioned genes, and the results indicate that the production of terpenoids might be a drought stress-dependent and cultivar-dependent procedure.

Molecular cloning, functional characterization and expression of a drought inducible phenylalanine ammonia-lyase gene (ObPAL) from Ocimum basilicum L.

Phenylalanine ammonia-lyase (PAL) is a control point for branched phenylpropanoid and terpenoid pathways. It represents the first regulatory step to provide a metabolic flux to produce of the precursors needed for biosynthesizing main volatile phenylpropanoid compounds (methyleugenol and methylchavicol) in basil. It is crucial during the stage of the environmental and development stimulants. To obtain better knowledge of the biosynthesis of these phenylpropene compounds, characterization and cloning of Ocimum basilicum PAL (ObPAL) cDNA and its heterologous expression and enzyme activity were assessed. The almost full-length ObPAL was 2064 bp in size encoding a 687-amino-acid polypeptide with molecular weight of 74.642 kDa and theoretical pI of 8.62. Phylogenetic analysis revealed a significant evolutionary relatedness of ObPAL with the PAL sequence reported in different species of Lamiaceae. To further confirm its function, ObPAL was cloned into pET28a (+) vector and expressed in E. coli. The recombinant protein exhibited high PAL activity and could catalyze the L-Phe conversion to trans-cinnamic acid. Expression analysis of PAL gene showed that ObPAL manifested various transcription ratios exposed to drought stress. Overall, our results demonstrated the ObPAL regulation gene is possibly a mechanism dependent on cultivar and drought stress.

Water deficit stress fluctuates expression profiles of 4Cl, C3H, COMT, CVOMT and EOMT genes involved in the biosynthetic pathway of volatile phenylpropanoids alongside accumulation of methylchavicol and methyleugenol in different Iranian cultivars of basil.

Here, for the first time, the accumulation ratio of methylchavicol and methyleugenoland compounds together with the expression profiles of five critical genes (i.e., 4Cl, C3H, COMT, CVOMT and EOMT) in three Iranian cultivars of basil were assessed under water deficit stress at flowering stage. The highest value of methylchavicol was detected for Cul. 3 under severe stress (S3; 7.695µg/mg) alongside Cul. 2 under similar circumstances (S3; 4.133µg/mg), while regarding Cul. 1, no detectable amounts were acquired. Considering methyleugenol, Cul. 3 (0.396µg/mg; S0) followed by Cul. 1 (S3; 0.160µg/mg) were the capable plant samples in producing some detectable amounts of methyleugenol. Apart from some expectations, all the genes under study exhibited also different transcription ratios under deficit stress. Our results, overall, demonstrated that the regulation of the above-mentioned genes and production of methychavicol and methyleugenol seems to be a cultivar- and drought stress-dependent mechanism.

Structure-activity relationship studies of 4-methylcoumarin derivatives as anticancer agents.

CONTEXT: Cancer is a leading cause of death worldwide and novel chemotherapeutic agents with better efficacy and safety profiles are much needed. Coumarins are natural polyphenolic compounds with important pharmacological activities, which are present in many dietary plants and herbal remedies. OBJECTIVES: The objective of this study is to investigate natural and synthetic coumarin derivatives with considerable anticancer capacity against three human cancer cell lines. MATERIALS AND METHODS: We synthesized 27 coumarin derivatives (mostly having 4-methyl moiety) and examined their cytotoxic effect on three human cancer cell lines, K562 (chronic myelogenous leukemia), LS180 (colon adenocarcinoma), and MCF-7 (breast adenocarcinoma) by MTT reduction assay. Screened compounds included 7-hydroxy-4-methylcoumarins (7-HMCs), 7-acetoxy-4-methylcoumarins (7-AMCs), and different dihydroxy-4-methylcoumarin (DHMC) and diacetoxy-4-methylcoumarin (DAMC) derivatives. Some compounds with methoxy, amine, and bromine substitutions were also examined. RESULTS: 7,8-DHMCs bearing alkyl groups at C3 position were the most effective subgroup, and of which, the most potent is compound 11, with an n-decyl chain at C3, which had IC50 values of 42.4, 25.2, and 25.1 µM against K562, LS180, and MCF-7 cells, respectively. The second most active subgroup was 7,8-DAMCs containing ethoxycarbonylmethyl and ethoxycarbonylethyl moieties at C3 position. Compound 27 (6-bromo-4-bromomethyl-7-hydroxycoumarin), the only derivative containing bromine also showed reasonable cytotoxic activities (IC50 range: 32.7-45.8 µM). DISCUSSION AND CONCLUSION: This structure-activity relationship (SAR) study of 4-methylcoumarins shows that further investigation of these derivatives may lead to the discovery of novel anticancer agents.