Biochimica et biophysica acta. General subjects最新文献

TOE1, also known as hCaf1z, belongs to the DEDD superfamily of deadenylases and a newly identified isoenzyme of hCaf1 deadenylases. Previous research has demonstrated that TOE1 has deadenylase activity, which can catalyze the degradation of poly(A) substrates and interact with hCcr4d to form the unconventional human Ccr4-Caf1 deadenylase complex. Our recent research indicates that hCaf1a and hCaf1b isoenzymes, highly expressed in gastric cancer, promote gastric cancer cell proliferation and tumorigenicity via modulating cell cycle progression. However, no studies have yet explored the relationship between TOE1 deadenylase and tumor development. In our study, we systematically investigated the functions and mechanisms of TOE1 in gastric cancer progression. Our findings revealed that overexpression of TOE1 inhibited gastric cancer cell proliferation, invasion and migration, promoted cell apoptosis, and led to cell cycle arrest in G0/G1 phase, while TOE1 knockdown had the opposite biological effects on these processes in gastric cancer cells. Further results indicated that TOE1 suppressed gastric cancer progression by inhibiting EMT process and MMPs expression. Moreover, our study clarified that TOE1 blocked gastric cancer cell cycle progression by up-regulating the expression level of the key cell cycle factors p21 and p53 through different regulatory mechanisms. Specifically, TOE1 up-regulated p53 expression by enhancing p53 promoter activity, and up-regulated p21 expression by enhancing p21 mRNA stability. Collectively, our findings first contribute to further elucidating the molecular mechanisms by which TOE1 participates in the regulation of gastric cancer progression, and are expected to provide a theoretical basis for diagnosis and targeted treatment of gastric cancer.

BMY 7378 is a multitarget drug primarily known for its selective antagonism of α_1D-adrenoceptors (α_1D-AR), exhibiting both hypotensive effects and the ability to prevent or reverse angiotensin II-induced vascular hypertrophy. Notably, BMY 7378 contains a phenylpiperazine moiety, a structural feature associated with angiotensin-converting enzyme (ACE) inhibition. This study aimed to investigate ACE inhibition as a potential pharmacological mechanism of BMY 7378. Using an in silico approach we predicted BMY 7378 interactions with the ACE active site, followed by in vitro activity assays. Additionally, ACE protein expression in the heart was analyzed following four weeks of BMY 7378 treatment in 7-8-month-old spontaneously hypertensive rats (SHR). All assays were benchmarked against captopril, a standard ACE inhibitor. In silico results showed that BMY 7378 binds to the ACE active site, though with reduced interaction with Zn701 (73.7 % compared to captopril), likely due to the pKa of its amino group. The inhibitory concentration 50 (IC₅₀) for BMY 7378 was 136 μM, lower than other reported phenylpiperazine derivatives. Furthermore, BMY 7378 significantly increased ACE expression in the hearts of SHR, with an increase of 8.5-fold compared to captopril. In conclusion, BMY 7378 exhibits dual activity as an α_1D-AR antagonist and an ACE inhibitor, making it a promising pharmacological tool for investigating and potentially treating hypertension and its associated cardiovascular complications.

Background: In earlier communications we reported about nanonization of the antibiotic tetracycline (Tet) by entrapping it within the biocompatible and highly membrane penetrating nano-carrier molecule - calcium phosphate nanoparticle (CPNP). The synthesized Tet-CPNP killed different Tet-resistant bacteria in vitro as well as in vivo (in mice). Moreover, such nanonized tetracycline had bactericidal mode of action, in contrast to bacteriostatic mode of action of bulk tetracycline. The present study unveils the molecular mechanism of action of Tet-CPNP.

Methods: This study was conducted to investigate the mode of interaction of Tet-CPNP/Tet with intact 70S bacterial ribosome by the techniques of spectrophotometry, spectrofluorimetry, circular dichroism, gel electrophoresis and transmission electron microscopy.

Results: Experimental observations revealed that (i) binding affinity of Tet-CPNP was higher than that of only tetracycline with ribosome and (ii) binding of Tet-CPNP, but not of tetracycline, loosened ribosome conformation, finally disrupting and degrading ribosome.

Conclusion: Bactericidal action of Tet-CPNP was rooted from degradation of cellular ribosomes and thereby blockage of protein translation phenomenon. Therefore, the problem of obsolescence of tetracycline, a cheap, first-generation, broad-spectrum antibiotic, due to generation of huge tetracycline-resistant bacteria, can be removed by the Tet-CPNP.

Background: N-Glycan branching modulates the diversity of protein functions. β1,4-N-acetylglucosaminyltransferase III (GnT-III or MGAT3) produces a unique GlcNAc branch, "bisecting GlcNAc", in N-glycans, and is involved in Alzheimer's disease and cancer. However, the 3D structure and catalytic mechanism of GnT-III are unclear. According to AlphaFold-based structure prediction, GnT-III likely contains two putative disordered segments, a long middle loop (Loop) and a C-terminal tail (Tail). We hypothesized that these segments play important roles in regulating the activity or intracellular behaviors of GnT-III.

Methods: We expressed wild-type GnT-III (GnT-III-WT), GnT-III-Loop- and -Tail-deletion mutants in cells. Their in vitro catalytic activity and glycan biosynthesis in cells were examined using high-performance liquid chromatography, UDP-Glo glycosyltransferase assays, and glycomic analysis. Subcellular localization of WT and GnT-III mutants was investigated by immunostaining, and degradation rate and secretion were also examined.

Results: The Loop-deletion mutant had higher in vitro and in cellulo activity than GnT-III-WT, indicating that Loop suppresses catalytic activity. In contrast, the Tail-deletion mutant showed weaker activity, increased ER localization, and faster degradation than GnT-III-WT, indicating that Tail is required for proper folding. In addition, deletion of Loop led to aberrant shedding of GnT-III, indicating that Loop contains the cleavage site or regulates GnT-III shedding.

Conclusions: Loop and Tail of GnT-III play important roles in catalytic activity, folding and shedding.

General significance: Our results provide further understanding of the catalysis and shedding mechanisms of GnT-III and can help in the development of methods for modifying the levels of bisecting GlcNAc on glycoproteins and in cells.

Background: Diabetic wounds, a significant complication of Type 2 Diabetes Mellitus (T2DM), face delayed healing due to impaired inflammation, angiogenesis, and collagen synthesis. This study explores Febuxostat, a xanthine oxidase inhibitor for its therapeutic potential in wound healing. Combining computational approaches and in-vitro assays, the study evaluates its effects on key wound healing pathways, cell viability, migration.

Methodology: The potential of Febuxostat in diabetic wound healing was studied using in-silico tools for Molecular docking and ADMET profiling, alongside Molecular dynamics (MD) simulations. Toxicity was assessed with OSIRIS Explorer, and biological activity was predicted using the PASS tool. In-vitro MTT and scratch assays on L929 cells further validated cytotoxicity and wound healing efficacy.

Results: Docking analysis revealed strong binding affinities to key wound healing targets, including VEGF (-9.11 kcal/mol) and NFKβ (-8.62 kcal/mol). Pharmacokinetic studies highlighted favorable skin permeability, supporting topical applications. Toxicity predictions indicated a safe profile. Molecular dynamics simulations demonstrated stable protein-ligand complexes, particularly with VEGF. Cytotoxicity studies on L929 cells revealed an IC₅₀ of 6.08 μM and the scratch assay demonstrated significant wound healing activity, highlighting its effectiveness in promoting cell migration and closure.

Conclusion: Febuxostat shows remarkable potential in enhancing diabetic wound healing by promoting cell migration, targeting wound-healing proteins, as demonstrated through in-silico and in-vitro studies. This drug is poised to effectively treat diabetic wounds, accelerating healing and reducing complications. Rigorous pre-clinical and clinical evaluations are essential to validate its safety, efficacy, and therapeutic potential.

Peptidyl prolyl cis/trans isomerases (PPIases), a ubiquitously distributed superfamily of enzymes, associated with signal transduction, trafficking, assembly, biofilm formation, stress tolerance, cell cycle regulation, gene expression and tissue regeneration, is a key regulator of metabolic disorders and microbial virulence. This review assumes an integrative approach, to provide a holistic overview of the structural and functional diversity of PPIases, examining their conformational dynamics, cellular distribution, and physiological significance. We explore their intricate involvement in cellular processes and virulence modulation in both eukaryotic and prokaryotic systems. Additionally, we evaluate the potential of these molecular chaperones as drug targets and vaccine candidates, emphasizing their relevance in therapeutic development. By synthesizing recent findings and providing a broader perspective on these proteins, this review aims to enhance our understanding of their multifaceted roles in biology and their potential applications in medicine.

The interaction of drugs with DNA is crucial for understanding their mechanism of action, particularly in the context of gene expression regulation. Erdafitinib (EDB), a pan-FGFR (fibroblast growth factor receptor) inhibitor approved by the FDA, is a potent anticancer agent used primarily in the treatment of urothelial carcinoma. In this study, the binding interaction between EDB and calf thymus DNA (ctDNA) was assessed using molecular docking, UV-absorption spectroscopy, fluorescence spectroscopy, and circular dichroism (CD) spectroscopy. The absorption spectra indicated a hypochromic effect when EDB was combined with ctDNA. The binding constant (K_a) of EDB-ctDNA complex was calculated as 7.84 × 10³ M^-1, corresponds to a free energy change (ΔG) value of approximately -5.06 kcal/mol, indicating a moderate binding affinity. Fluorometric analysis revealed a static binding mechanism in the ground state, with a bimolecular enhancement constant (K_B) of 7.56 × 10¹¹ M^-1. Displacement experiments demonstrated that EDB preferentially binds to the minor groove of ctDNA, with a Ksv value of 5.14 × 10⁴ M^-1. Further, KI quenching and CD spectroscopy confirmed the minor groove binding mode, which was associated with a decrease in the T_m from 68.28 °C to 65.84 °C, reflecting a destabilizing effect on DNA helix. Molecular docking supported these findings, showing that EDB exhibits a strong affinity for the minor groove of ctDNA and hydrogen bonding and Vander Waal interactions are the major forces involved in the binding. These results suggest that EDB primarily binds to the minor groove of ctDNA, which may play a role in its anticancer activity.

Background: Multidrug resistance (MDR) poses a significant obstacle to developing chemotherapeutic treatments. In previous studies using a traditional model of adriamycin resistance (ADR) with K562 cells, we demonstrated that N-acetylglucosaminyltransferase III (GnT-III) expression negatively regulates chemoresistance. Additionally, we observed that fucosylation levels were increased in the ADR cells.

Method: Fucosylation levels were determined using lectin blot, western blot, and flow cytometry. Gene expression levels were analyzed via qPCR. We generated a FUT4 knockout (KO) ADR cell line using CRISPR/Cas9 technology. Cytotoxicity and drug efflux assays were conducted to evaluate chemotherapy tolerance.

Results: The expression levels of FUT4 and its products, the Le^X antigens, were significantly upregulated in the ADR cells compared to the parental K562 cells. The FUT4 KO reduced the elevated levels of P-glycoprotein (P-gp) found in ADR cells and exhibited increased sensitivity to chemotherapeutic drugs. Furthermore, restoring FUT4 expression in the KO cells effectively reversed P-gp expression, drug efflux, and chemoresistance. Given the critical role of the NF-κB pathway in P-gp expression, we investigated NF-κB signaling and found that the phosphorylation levels of p65 were significantly increased in the ADR cells but were downregulated in the FUT4 KO cells. Furthermore, the restoration of FUT4 rescued the phosphorylation levels of p65.

Conclusions: FUT4 specifically upregulates P-gp expression related to chemoresistance through the NF-κB signaling pathway.

General significance: This study highlights the importance of FUT4 in chemoresistance and suggests it may serve as a promising target for combating MDR.

Background: Salinity stress affects plant growth, development, biomass, yield, as well as their survival. A series of signaling cascade is activated to cope the deleterious effect of salinity stress. Cytokinins are known for their regulatory roles from cell growth and expansion to abiotic stress signaling. Two component system (TCS) are important multistep phosphorelay signal transduction machinery converging cytokinin, ethylene and light signal transduction pathways together. Plant TCS comprises of histidine kinases, phosho-transfer proteins and response regulators. Histidine kinases perceive the signal and relay it to response regulator via histidine containing phosphor-transfer proteins.

Scope of review: Response regulators are one of the major and diverse component of TCS system which have been extensively studied for their role in plant growth, development and circadian rhythm. However, knowledge of their regulatory role in abiotic stress signaling is limited. This mini-review specifically focus on role of response regulators in salinity stress signaling.

Major conclusion: Response regulators is the divergent node of TCS machinery, where cross-talks with other stress-mediated, phytohormone-mediated, as well as, light-mediated signaling pathways ensues. Studies from past few years have established central role of response regulators in salinity stress, however, the detailed mechanism of their actions need to be studied further.

General significance: Response regulators act as both negative as well as positive regulator of salinity and cytokinin signaling, making it an excellent target to increase crop yield as well as stress tolerance capabilities.