The Lamiaceae family comprises over 7.200 medicinal plant species and represents a rich source of biologically active compounds. Lamium species have attracted increasing scientific interest due to their diverse bioactive properties and therapeutic potential. This review synthesizes the current evidence on the antioxidant, anticancer, and antimicrobial mechanisms associated with Lamium species and related Lamiaceae members. The therapeutic effects of these plants are largely attributed to their abundant secondary metabolites, including flavonoids and essential oils. As antioxidants, these compounds mitigate oxidative stress through free radical scavenging and redox modulation. The review highlights the cytotoxic effects observed in prostate, lung, and breast cancer cell lines, with key mechanisms involving apoptosis induction and reduced cell viability. Lamiaceae extracts demonstrate significant antimicrobial activity against multidrug-resistant pathogens by disrupting bacterial membrane integrity and inhibiting essential microbial enzymes. Overall, the bioactive profiles of Lamium species underscore their potential as promising candidates for natural product–based drug discovery and future clinical applications.
Throughout their life cycle, plants are capable of forming organs by differentiating from special tissues called meristems. The shoot apical meristem (SAM) produces all above ground organs, such as leaves, axillary shoots, and flowers, through the continuous regeneration of stem cells. A delicate balance exists between stem cell regeneration and differentiation. Stem cell homeostasis in the SAM is maintained by a precise and dynamic negative feedback loop established through WUSCHEL and CLAVATA signalling. There is an important correlation between WUS, which preserves stem cell identity, and CLV, which inhibits meristem overgrowth, in plant development. While the WUS protein moves upward, activating CLV3 expression, the CLV3 signalling protein moves downward, suppressing WUS, creating a dynamic equilibrium. The WUS-CLV feedback cycle contributes to the morphologic integrity and preservation of the plant’s vegetative and generative developmental stages. This review aims to clarify the genetic regulatory mechanisms that play a role in maintaining stem cell identity and limiting meristem size during plant development and to evaluate the functional significance of the WUS-CLV interaction in SAM. In addition to its primary role in developmental morphogenesis, this pathway holds agronomic importance in terms of organ number and related yield parameter.
Objective: Gastric ulcers are a major public health concern caused by the disrupted balance between protective mechanisms and harmful factors affecting the gastric mucosa. In this context, royal jelly (RJ), a natural secretion produced by worker honey bees, has been assessed for its possible ulcer-healing effects in vitro. Materials and Methods: An in vitro gastric ulcer model was established using a bile salt, sodium taurocholate (NaT), in the human gastric epithelial cell line (AGS). Non-toxic concentrations of RJ and an injurious dose of NaT were determined using the MTT assay. Cells were first exposed to NaT (15 mM for 1 h) and then treated with RJ (25 mu g/mL for 48 h). Cell viability, apoptotic markers, inflammatory mediators, and oxidative stress parameters, including intracellular reactive oxygen species (ROS), lipid peroxidation, and protein carbonyl levels, were evaluated. Results: NaT decreased cell viability and markedly induced apoptotic, inflammatory, and oxidative responses. RJ treatment improved cell viability, reduced pro-apoptotic markers (cleaved caspase-3 levels, Bax/Bcl-2 ratio), downregulated pro-inflammatory cytokines NF-kappa B p65, TNF-alpha, IL-1 beta, and IL-6, upregulated anti-inflammatory cytokine IL-10, and minimised ROS generation, lipid peroxidation, and protein carbonylation. RJ alone was non-toxic and increased IL-10 levels without provoking inflammation. Conclusion: RJ exerts cytoprotective effects against bile salt-induced gastric epithelial damage via antiapoptotic, anti-inflammatory, and antioxidant mechanisms, supporting its potential as a safe natural candidate for gastric mucosal protection and justifying further in vivo and clinical investigations.
Objective: Valproic acid (VPA) and its salts are psychotropic medications that are widely used to treat neurological diseases, such as epilepsy, migraine, and neuropathic pain, as well as psychiatric disorders, such as schizophrenia, addiction, and bipolar mood disorder. Moringa extract contains antioxidant, antimicrobial, antidiabetic, cardioprotective, and anti-inflammatory compounds that may support human health.Materials and Methods: This study aimed to investigate the possible protective effects of Moringa hydroalcoholic extract (MOL) against sodium valproate (SVP)-induced damage to brain glycoproteins and hydroxyproline. Female rats (Sprague Dawley) were randomly assigned to the following four groups: control, MOL, SVP, and SVP + MOL. SVP and MOL were administered at doses of 500 and 300 mg/kg body weight, respectively, for 15 days. Rats were sacrificed on day 16. The collection and subsequent homogenisation of brain tissue was conducted using a physiological saline solution at a ratio of 10% (weight/volume). The levels of sialic acid, hexose, hexosamine, fucose, and hydroxyproline were determined in the brain tissue.Results: Statistical analysis revealed a significant increase in glycoprotein and hydroxyproline levels in the brain tissue of rats treated with SVP. The administration of a hydroalcoholic extract of MOL to rats treated with SVP resulted in the recovery of these alterations, thus preventing or reducing brain damage.Conclusion: The hydroalcoholic extract of Moringa exerts an attenuating effect on brain damage associated with increased glycoprotein and hydroxyproline levels caused by SVP.
Objective: Sala lauak, a traditional fermented fish snack from West Sumatra, Indonesia, is a potential source of probiotic LAB. Information regarding LAB associated with this product remains limited. The spontaneous fermentation of salted fish under high-salt conditions selectively favours stress-adapted microorganisms. It is therefore hypothesised that LAB from sala lauak possess probiotic-related traits, including tolerance to acidic and bile conditions and antimicrobial activity against pathogenic bacteria. This study aimed to isolate and characterise LAB from sala lauak and evaluate its probiotic potential. Materials and Methods: Three isolates (SLA, SLB, and SLC) were purified and characterised based on morphological, biochemical, physiological, and molecular analyses. Tolerance to acid and bile salts was evaluated to assess gastrointestinal stress resistance. Antimicrobial activity was determined using inhibition zone assays against Escherichia coli O157, Salmonella sp., Staphylococcus aureus ATCC 25923, and Listeria monocytogenes. Molecular identification was performed using 16S rRNA gene sequencing. Results: All isolates were gram-positive, catalase-negative, rod-shaped bacteria with homofermentative metabolism. Isolate SLC showed the best overall gastrointestinal tolerance, with survival rates of 65.84 +/- 10.60% at pH 3 and 74.91 +/- 1.72% in 0.3% bile salts, and the strongest antimicrobial activity, producing inhibition zones of 29.10 +/- 0.99 mm against E. coli O157, 21.80 +/- 0.95 mm against Salmonella sp., 22.81 +/- 0.84 mm against S. aureus ATCC 25923, and 26.58 +/- 0.94 mm against L. monocytogenes. Molecular analysis identified all isolates as Weissella cibaria. Conclusion: W. cibaria SLC exhibited strong acid-bile tolerance and broad antimicrobial activity, highlighting its potential as a natural biopreservative and functional starter culture from sala lauak.
Objective: Bisphenol A (BPA) is an industrial chemical that disrupts the endocrine system and impairs the oxidant-antioxidant balance. This study investigates the effects of Panax ginseng (PxG) on liver and kidney damage induced by BPA.Materials and Methods: Rats were divided into the control, control+PxG, BPA, and BPA+PxG groups. BPA (50 mg/kg) was orally administered to the BPA groups. PxG-treated groups were orally administered 100 mg/kg PxG five times per week for six weeks. At the end of the six weeks, liver and kidney tissue samples were taken for oxidative stress determination.Results: BPA administration significantly increased oxidative stress. PxG administration decreased BPA-induced oxidative stress, as evidenced by a reduction in lipid peroxidation and restoration of antioxidant enzyme activity in liver and kidney tissues.Conclusion: PxG exhibits hepatoprotective and nephroprotective effects. Considering the widespread environmental exposure to BPA, PxG may represent a promising therapeutic agent for preventing BPA- induced hepatic and renal injury.
Objective: Global food security and ecosystem stability are under constant threat due to agricultural soil contamination driven by cadmium (Cd) and other heavy metals. Therefore, identification of tolerant and high-biomass-producing plant species is critical for sustainable phytoremediation. Materials and Methods: Ensete ventricosum (Welw.) Cheesman (wild banana) plants were grown in soil culture pots under controlled phytotron conditions and treated with different Cd concentrations (0, 10, 20 and 40 & micro;M) for 270 days. The accumulation of Cd, long-term morphological changes, physiological stress responses, and phytoremediation potential of E. ventricosum were evaluated. Results: Inductively coupled plasma optical emission spectroscopy (ICP-OES) revealed that Cd accumulation in plant organs was found in the order of root > corm > stem > leaf. Decreased Cd accumulation in corms with increased soil Cd shows a physiological barrier limiting metal translocation into storage organs. Plants exhibited a hormetic growth response when exposed to low Cd concentrations (10-20 & micro;M). E. ventricosum employed a tissue-specific and dose-dependent antioxidant defence strategy through the upregulation of catalase, guaiacol peroxidase, and ascorbate peroxidase activities to scavenge reactive oxygen species. In addition, the increased level of total soluble proteins in roots also reflects the synthesis of stress response proteins. Conclusion: E. ventricosum tolerates extended Cd exposure through coordinated enzymatic detoxification and morphological plasticity. E. ventricosum is a promising candidate for phytostabilization given its high plant biomass and high root Cd accumulation in Cd-contaminated soils.
Type 2 diabetes mellitus (T2DM) is often complicated by oxidative stress, lipid abnormalities, and a high cardiovascular disease burden. This review aims to examine the effects of paraoxonase 1 (PON1) gene polymorphisms and its antioxidant activity in relation to high-density lipoproteins and the risk of cardiovascular diseases, such as atherosclerosis and coronary artery disease, in patients with T2DM. The published literature was comprehensively reviewed, focusing on studies exploring genetic variants of PON1, enzymatic activity levels, ethnic distribution, and their interaction with metabolic and environmental factors in diabetic populations. The findings consistently showed that reduced PON1 activity is a major determinant of vascular risk in T2DM. Genetic variations alter enzyme function: the R allele of Q192R enhances paraoxon hydrolysis but reduces the protection of low-density lipoproteins, whereas the M allele of L55M decreases protein stability and circulating concentration. These effects vary across populations, with significant ethnic differences in allele prevalence. Beyond genetics, lifestyle and dietary factors, such as intake of saturated fats or polyphenol-rich foods, modulate PON1 activity, highlighting important gene-environment interactions. PON1 polymorphisms play a pivotal role in shaping cardiovascular vulnerability in patients with T2DM. Measuring enzyme activity may serve as a more accurate predictor of cardiovascular disease risk than genotyping alone because it reflects both inherited and environmental influences. Integrating genetic profiling with personalised dietary, lifestyle, and therapeutic interventions may lead to more precise cardiovascular risk assessment and improved outcomes for patients with diabetes. This approach offers a promising direction for future research and clinical practice.
Objective: Inhibitors of the ataxia-telangiectasia mutated (ATM) kinase are utilised to enhance the genotoxicity of chemotherapeutic agents. The two widely used ATM inhibitors (ATMi)-KU-55933 and KU-60019 are also recognised as autophagy inhibitors. Suppression of late-stage autophagy can promote the secretion of extracellularvesicles (EVs), and cancer-derived EVs are implicated in disease progression and metastasis. Therefore, this study aimed to elucidate the effects of these ATMi on EV secretion. Materials and Methods: The effects of ATMi on autophagy were studied using immunoblotting and microscopy in HeLa cells. EVs from treated cells were isolated via differential centrifugation and analyzed by immunoblotting and dynamic light scattering. The cytotoxicity of the treatments was evaluated using flow cytometry. Results: KU-60019 and KU-55933 caused LC3B-II and p62 accumulation in HeLa cells, as well as the secretion of large and small EVs containing both autophagy (LC3B-II, p62) and EV markers (CD63 and syntenin). The addition of etoposide to treatment with KU-60019 synergistically enhanced EV secretion and further increased LC3B-II accumulation in cell lysates, while reducing p62 accumulation. Nevertheless, compared to treatmentwith etoposide alone, the combined treatment resulted in higher levels of p62 and reduced gamma H2AX ubiquitination, which concurs with previous reports on the effects of p62 accumulation on the DDR. Conclusion: Our results reveal the potency of ATMi as EV inducers and the interplay between ATMi and etoposide in the regulation of EV secretion. The potential impact of ATMi-induced EVs on the tumour microenvironment should be considered in future trials.
Objective: Antibiotic resistance has become a global health issue, making it difficult to manage treatable infections. This study aimed to investigate the development of resistance in Klebsiella pneumoniae exposure to antibiotic diffusion gradients generated by disk diffusion. The emergence of cross-resistance and the impact of this resistance on biofilm formation are further evaluated. Materials and Methods: K. pneumoniae was exposed to antibiotic diffusion gradients using Oxoid disks (36 & micro;g) to select resistant mutants of piperacillin-tazobactam (TZP), leading to the development of a resistant strain (P-3). Comparative analyses of antibiotic susceptibility (TZP, amoxicillin/clavulanic acid [AMC], cefixime [CFM], and trimethoprim-sulfamethoxazole [SXT]) and biofilm formation were conducted between the parental (P-0) and resistant (P-3) strains. Biofilm production was evaluated under varying temperatures (37 degrees C and 45 degrees C) and glucose concentrations (0%-2.5%). Results: Resistance to TZP increased significantly in P-3, with moderate cross-resistance observed against AMC and SXT (p<0.05). Although the strain was susceptible to most antibiotics at baseline (P-0), it exhibited primary resistance to CFM in both P-0 and P-3 stages. Biofilm formation was higher at 37 degrees C than at 45 degrees C under the tested conditions. Increasing the glucose concentration from 0 % to 2.5% reduced biofilm production by nearly 40%. However, no direct correlation was found between antibiotic resistance development and biofilm-forming capacity. Conclusion: Selection through antibiotic gradients can induce resistance and cross-resistance in K. pneumoniae, but does not necessarily enhance biofilm production. Environmental parameters, particularly temperature and nutrient availability, and particularly glucose concentration, are more influential in modulating biofilm formation. Future molecular characterisation of beta-lactamase and biofilm-associated genes are essential for elucidating the underlying mechanisms.
Objective: This study demonstrated the successful application of multimodal chromatography (MMC) as an alternative to protein A chromatography for the purification of mAbs with anti-vascular endothelial growth factor properties. Materials and Methods: PROchievA and mAbSelect PrismA resins were used for protein A chromatographic purification, while Capto MMC and Nuvia cPrime resins were used for multimodal chromatographic purification. Each purification study was conducted as the initial capture step for the mAb produced in the Chinese hamster ovary cell culture. The experimental designs for the purification steps were developed using Modde 13 software, and the resulting data were analysed with the assistance of the same software. Results: Purification steps using protein A resins resulted in yields of approximately 87.2%-93%, whereas yields obtained with multimodal resins were approximately57.8%-65.2%. The multimodal purification step enabled the production of a monoclonal antibody (mAb) with a higher purity compared to the protein A purification step. It also showed superior performance compared to protein A resins in the removal of acidic and basic variants, achieving approximately 10% greater reduction of these variants in a single step and enabling the production of a purer mAb product. Conclusion: This study highlighted the potential of MMC to enhance the purity of mAbs with greater efficiency in removing charge variants. The purified mAbs were confirmed based on their mass values using the sodium dodecyl sulphate-polyacrylamide gel electrophoresis.
To disassemble the mechanisms of action of antipathogenic probiotics from Lactobacillus and Bifidobacterium genera, specifically competitive exclusion, bacteriocin production, short-chain fatty acid release, and immune modulation, to inform the development of microbiome-sparing compounds for multidrug-resistant infections. We conducted a systematic literature search across PubMed, Web of Science, Medline, and Google Scholar, identifying 104 pertinent articles published between 2018 and 2025. These studies were analysed to elucidate the antimicrobial mechanisms of probiotic-derived molecules and their potential for drug development. The analysis of 104 articles revealed that probiotic-derived molecules, including bacteriocins and short-chain fatty acids, create an adaptive anti-pathogen toolkit through competitive exclusion and immune modulation. These mechanisms effectively target MDR pathogens. However, translation to clinical settings remains complex due to inconsistent trial results and highly strain-specific therapeutic effects, underscoring the need for careful validation. Understanding the structural determinants of probiotic-derived molecules could enable the rational design of microbiome-compatible antivirulence drugs and bacteriocin mimetics that neutralise pathogens without harming the commensal microbiota. Harnessing these insights may extend the lifespan of existing antibiotics while advancing sustainable, ecosystem-friendly therapeutics for the post-antibiotic era.
Objective: This study investigated how stress factors (abscisic acid:ABA and heat shock) impact barley genotypes BOMI and its isogenic mutant, RISO-1508, by analyzing gene expression and transcriptomics.Materials and Methods: The impact of ABA on stress during barley germination was studied, followed by heat shock stress application. After treatment, total RNA was extracted, and cDNA libraries were created for transcriptomic analysis using Oxford Nanopore Technology. Genes with altered expression levels in heat-stressed barley varieties were identified. The most highly expressed DEGs were further examined through GO and KEGG enrichment analyses.Results: High concentrations of ABA almost completely inhibited barley germination and negatively impacted leaf growth, whereas root growth was less affected. This study is the first to assess the effects of heat stress on BOMI/RISO-1508 cells, revealing significant genetic changes under heat stress. Both genotypes showed similar protective mechanisms by significantly increasing heat shock proteins (HSPs) under heat stress, with BOMI uniquely upregulating five HSPs. A notable difference was observed in fatty acid and energy metabolism, indicating a more robust stress response in the BOMI group. Compared with BOMI, RISO-1508 showed greater downregulation of key photosynthetic genes, such as ATP synthase subunit beta and ribulose biphosphate carboxylase. Under HS, ABA receptor PYL4-like (LOC123424614) was significantly increased in BOMI, unlike in RISO-1508, indicating that BOMI has a distinct ABA stress response under HS.Conclusion: The findings indicate that environmental changes disrupt metabolic pathways in heat stress adaptation due to genotype variations, emphasizing the need to improve heat tolerance in barley.
Objective: The pursuit of an effective pharmaceutical agent to treat breast cancer has posed a significant challenge over decades. This work aims to study the therapeutic effect of the highly regarded medicinal plantAnnona muricata as an effective way to treat breast cancer. Materials and Methods: The leaf and fruit extracts of A. muricata were tested for their antioxidant, anti-inflammatory, anti-diabetic, antiproliferative potentials and their phytochemical composition was evaluated by liquid chromatography/mass spectroscopy (LC-MS). Results: The antioxidant evaluation indicated that the leaf and fruit extract had better effects than Gallic acid. The anti-inflammatory properties analysed by protein denaturation method showed 80% protection in the leaf extract and the haemolytic assay indicated a 90% protection, which was better than the Diclofenac standard. The leaf extracts proved to have excellent antidiabetic properties inhibiting alpha amylase to 76%, while the acarbose standard had an inhibition of 50%. Further cytotoxic effects of the leaf extract against breast cancer MCF-7 cells showed a dose-dependent inhibition of 88% at a concentration of 100 & micro;g while standard drug cisplatin showed an inhibition of 59% at a concentration of 50 & micro;g. However, the standard drug had no significant change beyond this concentration, which gave us a clear indication that the leaf extract had better cytotoxic activity. The LC-MS profiling showed the presence of various phytochemicals such as flavonoids, alkaloids, phenolics, terpenoids, and fatty acids, which may have contributed towards anti-inflammatory and anti-proliferative activity. Conclusion: The ability of A. muricata leaf and fruit extracts to prevent cancers could help prove the plant to be a potential alternative to synthetic drugs.
Objective: Hefianthemum germanicopofitanum Bornm., an endemic medicinal plant of T & uuml;rkiye belonging to the Cistaceae family, grows naturally in gypsum habitats characterised by drought, nutrient limitations, and soil toxicity. These severe abiotic conditions cause oxidative stress in plants by producing reactive oxygen species. Since the activation of antioxidantsystems is a typical plant response to stress, this study aimed to assess the antioxidant potential of H. germanicopofitanum by examining nutrient distribution and antioxidant enzyme activities in various tissues. Materials and Methods: Elemental content, along with the activities of guaiacol peroxidase (GPOX), superoxide dismutase (SOD), ascorbate peroxidase (APX), and catalase (CAT), as well as the levels of malondialdehyde (MDA) and hydrogen peroxide (H2O2), were measured in the roots, stems, and leaves. Results: The leaves exhibited higher concentrations of nitrogen (N), phosphorus (P), and potassium (K), whereas the roots contained greater amounts of calcium (Ca), magnesium (Mg), iron (Fe), manganese (Mn), and copper (Cu) ions. GPOX, APX activities and H2O2 were highest in the leaves, whereas CAT and MDA were most concentrated in the roots. The SOD activity was consistent across tissues. Conclusion: Enzyme activity was notably high in the roots and leaves, indicating the strong antioxidant capacity of the plant in adapting to stressful gypsum environments.
Objective: Plants rely on nucleotide excision repair to eliminate UV-induced pyrimidine dimers, which disrupt transcription and replication, thereby supporting growth and development under UV stress. In this process, endonucleases, the XPF-ERCC1 complex, and XPG in mammals make dual incisions around the damage site. In Arabidopsis thaliana, the XPF homolog UVH1 has been associated with several DNA repair pathways, but direct evidence for its role in the cleavage of damaged oligonucleotides during nucleotide excision repair has been limited. Materials and Methods: An excision assay was performed on UV-treated wild-type and uvh1 mutant Arabidopsis plants to examine the nucleotide excision repair activity. Phylogenetic analysis of XPF homologs was conducted across plant and other eukaryotic lineages. UVH1 transcript levels were analyzed in different developmental stages, tissue types, and stress conditions. Results: UVH1 is required for the excision of UV-induced pyrimidine dimers, which directly supports its role in the dual incision step of plant nucleotide excision repair. Phylogenetic analysis revealed that plant XPF homologs form a distinct, evolutionarily conserved lineage. UVH1 expression is regulated in a tissueand development-specific manner, responding dynamically to environmental stresses. Conclusion: This study provides direct evidence that UVH1 mediates dual incision during nucleotide excision repair in Arabidopsis, confirming its functional conservation with the animal XPF protein.
Objective: This study presents the first investigation of the biological activities of Camellia cattienensis extract, an endemic tea species in Vietnam. Materials and Methods: Leaves collected from Cat Tien National Park were subjected to methanol extrac tion and phytochemical screening, which confirmed the presence of alkaloids, coumarins, flavonoids, glycosides, saponins, polyphenols, polyuronoids, reducing sugars, and steroids. Results: The extract exhibited high levels of phenolics (1210.92 +/- 30.70 mg GAE/g) and flavonoids (744.40 +/- 24.31 mg QE/g), with catechin profiling identifying epicatechin, catechin, epigallocatechin gallate, and epicatechin gallate. Antibacterial activity was notable against Bacillus cereus (MIC=2.00 mg/mL; MBC=5.00 mg/mL), but absent against Gramnegative bacteria. Strong antioxidant potential was demon strated (DPPH EC50=3.27 +/- 0.12 & micro;g/mL; reducing power comparable to Trolox). Cytotoxicity assays revealed selective inhibition of MCF7 and K562 cells, with higher potency against K562 (IC50=63.05 +/- 2.83 & micro;g/mL, SI=3.04). No acute toxicity was observed, underscoring the therapeutic promise of C. cattienensis. Conclusion: C. cattienensis extract exhibited antioxidant and anticancer effects.
Objective: Burn injuries are a significant global health concern that are frequently intensified by infections. Fungal infections, especially those produced by Candida species, are a leading cause of morbidity and mortality in burn patients. This study investigated the prevalence, species distribution, and antifungal susceptibility of fungal pathogens in burn wound infections in Duhok, Iraq. Materials and Methods: From September 2024 to March 2025, 156 burn wound swabs were obtained from patients attending burn units in Duhok. Samples were cultured on Sabouraud dextrose agar and blood agar. Fungal isolates were identified using conventional methods and validated by PCR amplification and ITS region sequencing. Antifungal susceptibility testing was performed for fungal isolates. Results: Fungal infections were observed in 25.6% of cases (40/156), with Candida spp. accounting for 52.5% (21/40) of isolates. The predominant species was Candida afbicans (71.4%, 15/21), followed by Candida parapsifosis (23.8%, 5/21) and Candida tropicafis (4.7%, 1/21). The most common isolates were found in the leg (42.8%) and hand (28.5%), primarily caused by seconddegree burns (61.9%) and hot fluid (42.8%). Mortality among infected patients was 19.0%. Most Candida isolates were nystatinresistant, with variable sensitivity to other antifungals Conclusion: Candida species, specifically Candida afbicans, are the most prevalent pathogens in burn wound infections in Duhok. The early and accurate identification of fungal species, coupled with antifungal susceptibility profiling, is the key to optimising therapeutic strategies and improving patient outcomes.
Objective: In Agrobacterium tumefaciens, zinc allocation occurs through zinc chaperones, which are regulated by the Zur (Zinc uptake regulator) repressor. The putative COG0523-family chaperones atu4502 and atu3633 contain consensus Zur-binding sequences upstream of their start codons. The purpose of this study was to investigate their regulation and possible functions. Materials and Methods: Quantitative real-time PCR (qRT-PCR), translational promoter-fusion assays and DNase I footprinting were employed to determine the regulation. All phenotypic assays were carried out in AB minimal media. Results: qRT-PCR analyses revealed that atu4502 and atu3633, encoding novel members of the COG0523-family, are negatively regulated by Zur. DNase I footprinting indicated that Zur binds to consensus Zur box sequences overlapping-35 and-10 promoter sequences upstream of each gene. Translational promoter-fusion assays indicated that Zur boxes are necessary for transcriptional repression. Phenotypic assays showed that an atu4502 inactivation mutant is sensitive to the cell impermeable metal chelator, EDTA, whereas an atu3633 inactivation mutant is sensitive to the cell permeable metal chelator, TPEN, suggesting that the gene products may occupy different subcellular locations. Zinc supplementation was able to rescue both the EDTA sensitivity of atu4502::KM and the TPEN sensitivity of atu3633::GM, consistent with their proposed role as Zinc chaperones. Biofilm and EPS assays showed that in atu4502 and atu3633 inactivation mutants, biofilm and EPS levels were increased relative to wild-type. Conclusion: It was concluded that both atu4502 and atu3633 are regulated by Zur, and likely participate in zinc trafficking.
