Inhibition Of Protein Synthesis Research Articles

Synthesis and Evaluation of Novel 5‐O‐(4‐C‐Aminoalkyl‐β‐D‐ribofuranosyl) Apramycin Derivatives for the Inhibition of Gram‐Negative Pathogens Carrying the Aminoglycoside Phosphotransferase(3′)‐Ia Resistance Determinant

AbstractWe report the synthesis and evaluation of two new apramycin 5‐O‐β‐d‐ribofuranosides, or apralogs, carrying aminoalkyl branches at the ribofuranose 4‐position. This novel modification conveys excellent activity for the inhibition of protein synthesis by wild‐type bacterial ribosomes and correspondingly high antibacterial activity against several Gram‐negative pathogens. Notably, these new modifications overcome the reduction of antibacterial activity in other 2‐deoxystreptamine‐type aminoglycosides carrying a 5‐O‐ribofuranosyl moiety when challenged by the presence of an aminoglycoside phosphotransferase enzyme capable of acting on the ribose 5‐position.

Strategies for the Discovery of Oxazolidinone Antibacterial Agents: Development and Future Perspectives.

Oxazolidinones represent a significant class of synthetic bacterial protein synthesis inhibitors that are primarily effective against Gram-positive bacteria. The commercial success of linezolid, the first FDA-approved oxazolidinone antibiotic, has motivated researchers to develop more potent oxazolidinones by employing various drug development strategies to fight against antimicrobial resistance, some of which have shown promising results. Thus, this Perspective aims to discuss the strategies employed in constructing oxazolidinone-based antibacterial agents and summarize recent advances in discovering oxazolidinone antibiotics to provide valuable insights for potentially developing next-generation oxazolidinone antibacterial agents or other pharmaceuticals.

THU498 Establishing The Link Between Glucocorticoid Signaling And The Circadian Regulator DEC1 In Breast Cancer

Abstract Disclosure: M.I. Castro: None. W.S. Ybañez: None. P.D. Bagamasbad: None. Epidemiological studies have linked altered circadian rhythms due to irregular shift work, chronic jet lag, and heightened night-time light exposure to increased breast cancer (BCa) pathogenesis. Dysregulated glucocorticoid (GC; CORT) signaling in BCa may further promote chronodisruption by directly regulating the cyclic expression of clock genes found in the breast. The differentiated embryo-chondrocyte expressed gene 1 (DEC1; also known as STRA13/SHARP2/ BHLHB2) is a transcription factor involved in the regulation of circadian rhythmicity and tumor progression of several cancer types including the breast and associated with hormone receptor signaling in BCa. In this study, we explored the involvement of DEC1 in the crosstalk between GC signaling and the circadian axis in BCa. To establish DEC1 expression pattern in BCa tumor samples, we assessed publicly available RNA-seq datasets from the Genotype-Tissue Expression Project and found no significant difference in DEC1 expression in tumors when compared to normal tissue controls. However, when tumor samples were stratified into molecular subtypes based on transcriptomic data from the Cancer Genome Atlas, DEC1 expression was significantly downregulated in triple-negative BCa (TNBC) in comparison to luminal subtypes. This pattern of expression was reflected in the BCa cell lines where we observed higher DEC1 expression in the estrogen receptor-positive MCF7 cells relative to nontumorigenic triple-negative MCF10A cells and the highly aggressive TNBC MDA-MB-231 cell line. To determine if DEC1 is regulated by GCs in the breast, we treated non-tumorigenic breast epithelial MCF10A cells and the TNBC MDA-MB-231 cells with increasing concentrations of CORT and observed that DEC1 mRNA expression decreases in response to increasing concentrations of CORT. Treatment of MCF10A and MDA-MB-231 cells with GC receptor (GR)- specific agonist RU486 abolished the GC-dependent repression of DEC1 expression and this pattern of DEC1 expression persisted even in the presence of the protein synthesis inhibitor cycloheximide, suggesting that DEC1 is a direct transcriptional target of GR in mammary epithelial cells. Through in silico analysis of publicly available ChIP-sequencing datasets from BCa cell lines, we identified multiple CORT-responsive cis-regulatory elements in the DEC1 locus that could mediate the CORT-dependent repression of DEC1 expression. Lentiviral knockdown of DEC1 in three breast epithelial cell lines was generated and will be used in cancer hallmark assays on proliferation, apoptosis, survival, and migration to assess the functional role of DEC1 in BCa. Taken together, our data will contribute to understanding the combined effect of GC signaling and circadian gene regulation towards BCa progression. Presentation: Thursday, June 15, 2023

THU573 Paxillin Regulates Androgen Receptor In Granulosa Cells

Abstract Disclosure: A. Weidner: None. A. Roy: None. K. Vann: None. O. Astapova: None. Paxillin is a ubiquitously-expressed adaptor protein that is integral in focal adhesions and plays an active role in the regulation of cell motility and apoptosis. Recently, paxillin was shown to mediate pro-tumorigenic androgen signaling in prostate cancer cells, where it is necessary for rapid cytoplasmic kinase signaling triggered by activation of membrane-bound androgen receptor. We sought to investigate the relationship between paxillin and androgen receptor (AR) in granulosa cells, where androgen actions, apoptosis and focal adhesions are of known importance, but the role of paxillin is understudied. When paxillin expression was silenced using siRNA in human granulosa-derived KGN cells, AR protein expression was significantly reduced, while its RNA expression was unaffected. Similarly, in granulosa cell-specific paxillin knockout mice, AR protein expression in granulosa cells was reduced. Further, we found that AR and paxillin co-localize in focal adhesions in mouse primary granulosa cells and in KGN cells, and their interaction requires the activity of focal adhesion kinase. Disruption of focal adhesion with a focal adhesion kinase inhibitor also leads to reduced AR protein levels. To further explore the mechanisms by which paxillin regulates AR expression, we created paxillin-null KGN cell lines using CRISPR-Cas9 gene editing. While paxillin loss resulted in reduced AR protein expression early (days) in the gene-editing process, this effect disappeared after weeks of clonal expansion from selected single cells. Expanded paxillin-null KGN clones showed normal AR protein expression, but increased AR mRNA expression, suggesting an adaptive increase in AR gene transcription to compensate for reduced AR protein levels in cells that survived paxillin deletion. Studies of AR protein degradation using cycloheximide to block protein synthesis showed that the half-life of AR was reduced from 6.7 to 4.5 hours (approximately 33%) by paxillin deletion, indicating accelerated AR protein degradation in cells lacking paxillin. Based on our results, we hypothesize that paxillin brings AR into focal adhesions in granulosa cells, thereby sequestering it from proteasomal degradation in the cytoplasm. Thus, paxillin may be a novel target in the management of androgen-related disorders in women, such as polycystic ovary syndrome. Presentation Date: Thursday, June 15, 2023

YIV-818-A: a novel therapeutic agent in prostate cancer management through androgen receptor downregulation, glucocorticoid receptor inhibition, epigenetic regulation, and enhancement of apalutamide, darolutamide, and enzalutamide efficacy.

Introduction: Prostate cancer is the second leading cause of cancer death among men in the United States. Castration-Resistant Prostate Cancer (CRPC) often develops resistance to androgen deprivation therapy. Resistance in CRPC is often driven by AR variants and glucocorticoid receptor (GR). Thus, drugs that target both could be vital in overcoming resistance. Methods: Utilizing the STAR Drug Discovery Platform, three hundred medicinal plant extracts were examined across 25 signaling pathways to identify potential drug candidates. Effects of the botanical drug YIV-818-A, derived from optimized water extracts of Rubia cordifolia (R.C.), on Dihydrotestosterone (DHT) or Dexamethasone (DEX) induced luciferase activity were assessed in 22RV1 cells harboring the ARE luciferase reporter. Furthermore, the key active compounds in YIV-818-A were identified through activity guided purification. The inhibitory effects of YIV-818-A, RA-V, and RA-VII on AR and GR activities, their impact on AR target genes, and their roles in modifying epigenetic status were investigated. Finally, the synergistic effects of these compounds with established CRPC drugs were evaluated both in vitro and in vivo. Results: YIV-818-A was found to effectively inhibit DHT or DEX induced luciferase activity in 22RV1 cells. Deoxybouvardin (RA-V) was identified as the key active compound responsible for inhibiting AR and GR activities. Both YIV-818-A and RA-V, along with RA-VII, effectively downregulated AR and AR-V proteins through inhibiting protein synthesis, impacted the expression of AR target genes, and modified the epigenetic status by reducing levels of Bromodomain and Extra-Terminal proteins (Brd2/Brd4) and H3K27Ac. Furthermore, these compounds exhibited synergistic effects with apalutamide, darolutamide, or enzalutamide, and suppressed AR mediated luciferase activity of 22RV1 cells. Co-administration of YIV-818-A and enzalutamide led to a significant reduction of 22RV1 tumor growth in vivo. Different sources of R.C. had variable levels of RA-V, correlating with their potency in AR inhibition. Discussion: YIV-818-A, RA-V, and RA-VII show considerable promise in addressing drug resistance in CRPC by targeting both AR protein and GR function, along with modulation of vital epigenetic markers. Given the established safety profile of YIV-818-A, these findings suggest its potential as a chemopreventive agent and a robust anti-prostate cancer drug.

On the participation of adenosinergic receptors in the reconsolidation of spatial long-term memory in male rats.

To date, there is insufficient evidence to explain the role of adenosinergic receptors in the reconsolidation of long-term spatial memory. In this work, the role of the adenosinergic receptor family (A1, A2A, A2B, and A3) in this process has been elucidated. It was demonstrated that when infused bilaterally into the hippocampal CA1 region immediately after an early nonreinforced test session performed 24 h posttraining in the Morris water maze task, adenosine can cause anterograde amnesia for recent and late long-term spatial memory. This effect on spatial memory reconsolidation was blocked by A1 or A3 receptor antagonists and mimicked by A1 plus A3 receptor agonists, showing that this effect occurs through A1 and A3 receptors simultaneously. The A3 receptor alone participates only in the reconsolidation of late long-term spatial memory. When the memory to be reconsolidated was delayed (reactivation 5 d posttraining), the amnesic effect of adenosine became transient and did not occur in a test performed 5 d after the reactivation of the mnemonic trace. Finally, it has been shown that the amnesic effect of adenosine on spatial memory reconsolidation depends on the occurrence of protein degradation and that the amnesic effect of inhibition of protein synthesis on spatial memory reconsolidation is dependent on the activation of A3 receptors.

Comparative Proteomic Analysis of Two Wild Soybean (Glycine soja) Genotypes Reveals Positive Regulation of Saline-Alkaline Stress Tolerance by Tonoplast Transporters.

Soil saline-alkalization is a significant constraint for soybean production. Owing to higher genetic diversity of wild soybean, we compared the proteomic landscape of saline-alkaline stress-tolerant (SWBY032) and stress-sensitive (SWLJ092) wild soybean (Glycine soja) strains under saline and saline-alkaline stress. Out of 346 differentially expressed proteins (DEPs) specifically involved in saline-alkaline stress, 159 and 133 DEPs were identified in only SWLJ092 and SWBY032, respectively. Functional annotations revealed that more ribosome proteins were downregulated in SWLJ092, whereas more membrane transporters were upregulated in SWBY032. Moreover, protein-protein interaction analysis of 133 DEPs revealed that 14 protein-synthesis- and 2 TCA-cycle-related DEPs might alter saline-alkaline tolerance by affecting protein synthesis and amino acid metabolism. Furthermore, we confirmed G. soja tonoplast intrinsic protein (GsTIP2-1 and GsTIP2-2), inositol transporter (GsINT1), sucrose transport protein (GsSUC4), and autoinhibited Ca2+-ATPase (GsACA11) as tonoplast transporters can synergistically improve saline-alkaline tolerance in soybean, possibly by relieving the inhibition of protein synthesis and amino acid metabolism. Overall, our findings provided a foundation for molecular breeding of a saline-alkaline stress-tolerant soybean.

The Biological Action and Structural Characterization of Eryngitin 3 and 4, Ribotoxin-like Proteins from Pleurotus eryngii Fruiting Bodies.

Ribotoxin-like proteins (RL-Ps) are specific ribonucleases found in mushrooms that are able to cleave a single phosphodiester bond located in the sarcin-ricin loop (SRL) of the large rRNA. The cleaved SRL interacts differently with some ribosomal proteins (P-stalk). This action blocks protein synthesis because the damaged ribosomes are unable to interact with elongation factors. Here, the amino acid sequences of eryngitin 3 and 4, RL-Ps isolated from Pleurotus eryngii fruiting bodies, were determined to (i) obtain structural information on this specific ribonuclease family from edible mushrooms and (ii) explore the structural determinants which justify their different biological and antipathogenic activities. Indeed, eryngitin 3 exhibited higher toxicity with respect to eryngitin 4 against tumoral cell lines and model fungi. Structurally, eryngitin 3 and 4 consist of 132 amino acids, most of them identical and exhibiting a single free cysteinyl residue. The amino acidic differences between the two toxins are (i) an additional phenylalanyl residue at the N-terminus of eryngitin 3, not retrieved in eryngitin 4, and (ii) an additional arginyl residue at the C-terminus of eryngitin 4, not retrieved in eryngitin 3. The 3D models of eryngitins show slight differences at the N- and C-terminal regions. In particular, the positive electrostatic surface at the C-terminal of eryngitin 4 is due to the additional arginyl residue not retrieved in eryngitin 3. This additional positive charge could interfere with the binding to the SRL (substrate) or with some ribosomal proteins (P-stalk structure) during substrate recognition.

Phenomenological interpretations of the mechanism for the concentration-dependent positive effect of antibiotic lincomycin on Streptomyces coelicolor A3(2).

The antibiotic lincomycin binds to the 23S ribosomal RNA peptidyl transferase loop region to inhibit protein synthesis. However, lincomycin can also stimulate the growth and secondary metabolism of actinomycetes in a concentration-dependent manner. In Streptomyces coelicolor A3(2), lincomycin stimulates the production of the blue-pigmented antibiotic actinorhodin at concentrations below the minimum inhibitory concentration. To better understand the molecular mechanism underlying these concentration-dependent positive effects, this study investigated how the target molecule, the ribosome, undergoes dynamic changes in the presence of lincomycin and explored the ribosome-related factors involved. Lincomycin, at a concentration that stimulates actinorhodin production of S. coelicolor A3(2), could restore temporarily arrested ribosome function by utilizing ribosome-related proteins and translation factors, presumably under the control of the transcription factor WblC protein that confers intrinsic resistance to multiple translation-inhibiting antibiotics, to eventually produce stable and active ribosomes even during the late growth phase. This qualitatively and quantitatively positive ribosome alteration can be advantageous for producing actinorhodin biosynthetic enzymes. A series of gene expression and biochemical analyses revealed that lincomycin at the concentration that induces ribosomal stabilization in S. coelicolor A3(2) could influence the localization of the 20S proteasome-related proteins, resulting in reduced proteasome activity. These findings suggest that the functional analysis of 20S proteasome represents a potential pivotal challenge for understanding the molecular mechanism of ribosome stabilization induced by lincomycin. Therefore, as lincomycin can dynamically alter its target molecule, the ribosome, we discuss the future issues and prospects for an increased understanding of the concentration-dependent properties of antibiotics. IMPORTANCE Antibiotics were originally defined as chemical compounds produced by a microbe that inhibits the growth of other microbes. However, an unexplained effect of this is that a low concentration of antibiotics, such as those below the minimum inhibitory concentration, can positively affect microbial growth and metabolism. The secondary metabolic activation of streptomycetes in the presence of the translation-inhibiting antibiotic lincomycin illustrates the concentration-dependent positive effect of the antibiotic. The significance of this study is that the phenomenological interpretation of the molecular mechanism of the concentration-dependent positive effect of lincomycin in Streptomyces coelicolor A3(2) has provided novel insight into the possible role of antibiotics in making their target molecules stable and active with the assistance of various related factors that benefit their function. Further exploration of this idea would lead to an essential understanding of antibiotics, including why actinomycetes make them and their role in nature.

New perspective on mechanism in muscle toxicity of ochratoxin A: Model of juvenile grass carp (Ctenopharyngodon idella)

Ochratoxin A (OTA) is a common fungal toxin that pollutes raw materials of aquatic feeds (such as corn, soybean meal, and wheat). This study explored the effects of OTA through diet on muscle toxicity in juvenile grass carp (Ctenopharyngodon idella). The following results were obtained for the muscle. (1) With an increase in dietary OTA, the residue of OTA in muscle increased, muscle fiber diameter and density decreased, and even muscle fiber breakage. (2) OTA caused oxidative stress by downregulating GPx1 (a, b) and Trx via inhibited the PGC1-α/Nrf2 signaling pathway. (3) OTA exacerbated endoplasmic reticulum stress in the muscle by causing endoplasmic reticulum expansion (results of transmission electron microscopy) and upregulating the expression of GRP78, eIF2α, ATF6, PERK, and CHOP. (4) OTA reduced muscle fiber diameter by inhibiting protein synthesis (AKT, TOR, and S6K1) and promoting the mRNA expression of protein degradation-related genes (MURF1, MAFBX, and FoxO3a), as well as by reducing AKT and promoting the immunofluorescence expression of FoxO3. (5) OTA inhibits collagen deposition by downregulating TGF-β1, TGF-βR1, Smad2, Smad3, Smad4, CTGF, TIMP, PHD, and LOX mRNA expressions as well as the CTGF immunofluorescence expression. Moreover, based on the GSH and collagen content contents, the upper safe dose for OTA-induced toxicity was 963.6 and 1129.6 μg/kg diet, respectively. Using the example of OTA, our research has provided new insights that raise concerns about the quality of aquatic products by exploring muscle toxicity caused by mycotoxins.

Antifungal Activity of Ageritin, a Ribotoxin-like Protein from Cyclocybe aegerita Edible Mushroom, against Phytopathogenic Fungi.

Ageritin from poplar mushrooms is a specific endonuclease that hydrolyzes a single phosphodiester bond located in the sarcin-ricin loop (SRL) of the large rRNA, thereby blocking protein synthesis. Considering the possible biotechnological use of this enzyme, here we report its antifungal activity against virulent fungi affecting crops of economic interest. Our results show that ageritin (200 µg/plug; ~13.5 nmole) inhibits the growth of Botrytis cinerea (57%), Colletotrichum truncatum (42%), and Alternaria alternata (57%), when tested on potato dextrose agar plates. At the same time, no effect was observed against Trichoderma harzianum (a fungus promoting beneficial effects in plants). To verify whether the antifungal action of ageritin against B. cinerea and T. harzianum was due to ribosome damage, we tested ageritin in vitro on partially isolated B. cinerea and T. harzianum ribosomes. Interestingly, ageritin was able to release the Endo's fragment from both tested fungal ribosomes. We therefore decided to test the antifungal effect of ageritin on B. cinerea and T. harzianum using a different growth condition (liquid medium). Differently from the result in solid medium, ageritin can inhibit both B. cinerea and T. harzianum fungal growth in liquid medium in a concentration-dependent manner up to 35.7% and 38.7%, respectively, at the highest concentration tested (~200 µg/mL; 12 µM), and the analysis of RNA isolated from ageritin-treated cells revealed the presence of Endo's fragment, highlighting its ability to cross the fungal cell wall and reach the ribosomes. Overall, these data highlight that the efficacy of antifungal treatment to prevent or treat a potential fungal disease may depend not only on the fungal species but also on the conditions of toxin application.

Competition for calnexin binding regulates secretion and turnover of misfolded GPI-anchored proteins.

In mammalian cells, misfolded glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) are cleared out of the ER to the Golgi via a constitutive and a stress-inducible pathway called RESET. From the Golgi, misfolded GPI-APs transiently access the cell surface prior to rapid internalization for lysosomal degradation. What regulates the release of misfolded GPI-APs for RESET during steady-state conditions and how this release is accelerated during ER stress is unknown. Using mutants of prion protein or CD59 as model misfolded GPI-APs, we demonstrate that inducing calnexin degradation or upregulating calnexin-binding glycoprotein expression triggers the release of misfolded GPI-APs for RESET. Conversely, blocking protein synthesis dramatically inhibits the dissociation of misfolded GPI-APs from calnexin and subsequent turnover. We demonstrate an inverse correlation between newly synthesized calnexin substrates and RESET substrates that coimmunoprecipitate with calnexin. These findings implicate competition by newly synthesized substrates for association with calnexin as a key factor in regulating the release of misfolded GPI-APs from calnexin for turnover via the RESET pathway.

An Update on the Therapeutic Potential of Antimicrobial Peptides against Acinetobacter baumannii Infections.

The rise in antibiotic-resistant strains of clinically important pathogens is a major threat to global health. The World Health Organization (WHO) has recognized the urgent need to develop alternative treatments to address the growing list of priority pathogens. Antimicrobial peptides (AMPs) rank among the suggested options with proven activity and high potential to be developed into effective drugs. Many AMPs are naturally produced by living organisms protecting the host against pathogens as a part of their innate immunity. Mechanisms associated with AMP actions include cell membrane disruption, cell wall weakening, protein synthesis inhibition, and interference in nucleic acid dynamics, inducing apoptosis and necrosis. Acinetobacter baumannii is a critical pathogen, as severe clinical implications have developed from isolates resistant to current antibiotic treatments and conventional control procedures, such as UV light, disinfectants, and drying. Here, we review the natural AMPs representing primary candidates for new anti-A. baumannii drugs in post-antibiotic-era and present computational tools to develop the next generation of AMPs with greater microbicidal activity and reduced toxicity.

P10.28.B INTERMODEL VARIATION OF BBB PERMEABILITY AFFECTS CNS DRUG DELIVERY IN PDX MODELS OF GBM

Abstract BACKGROUND Little progress has been made in the development of effective new therapies for glioblastoma (GBM) the past decades. One of the major obstacles in identifying novel candidate drugs against GBM remains the blood-brain barrier (BBB) and the efflux transporters. Therefore, it is crucial to select drugs able to cross BBB and reach the tumor tissue in therapeutically effective concentrations. In current study, we evaluated the anti-glioma effect of the protein synthesis inhibitor omacetaxine mepesuccinate (OMA) in vitro and its CNS penetrating properties in vivo. MATERIAL AND METHODS The FDA-approved Oncology Drug Set II library was tested on 55 patient-derived GBM cell cultures. We designed a drug shortlisting pipeline combining efficacy data with pharmacodynamic and pharmacokinetic characteristics of each compound. We developed and characterized 3 orthotopic mouse PDX models in terms of brain vascular integrity, BBB permeability and expression of ABC transporters. The PK profile of omacetaxine was assessed in these orthotopic mouse PDX models. RESULTS In vitro, OMA revealed anti-tumor activity at IC50 values well-below reported Cmax plasma value (Cmax=43nM) in approximately 80% of GBM cultures. Unexpectedly, OMA does not downregulate the expression of its acknowledged target, ribosomal protein 3 (RPL3). In glioma cells while activation of caspase 3/7 activity is linked to its anti-tumor effect. OMA is a substrate for ABCB1 (P-glycoprotein) but not for ABCG2 (BCRP). In vivo, dose escalation showed safety of the drug and OMA was found to reach the brain tumor tissue in concentrations similar or higher to the reported IC50 values in vitro, depending on the PDX model used. No adverse reactions were noted. CONCLUSION OMA is a CNS penetrant drug that reaches the brain tumor tissue in all tested concentrations. The inter-model variation in accumulation of OMA is associated to the extent of BBB disruption of each PDX model. Additional in vivo experiments are ongoing to investigate the therapeutic effect of OMA and longer exposure using in vivo models.

Acute Oxidative Stress Can Paradoxically Suppress Human NRF2 Protein Synthesis by Inhibiting Global Protein Translation.

The NRF2 transcription factor is a master regulator of the cellular oxidant/electrophile response and a drug target for the prevention/treatment of chronic diseases. A major mechanism of NRF2 activation is its escape from rapid degradation, and newly synthesized NRF2 induces cytoprotective protein expression through its cognate antioxidant response elements (AREs). However, oxidative stress can also inhibit global protein translation, thereby potentially inhibiting NRF2 protein accumulation. H2O2 has been shown to be a relatively weak inducer of NRF2 in comparison with electrophiles. In the current study, we evaluated whether levels of H2O2 that activate the NRF2/ARE pathway inhibit NRF2 protein synthesis in HaCaT keratinocytes. A weak maximum induction was observed for H2O2 in comparison with electrophiles, both for NRF2 protein accumulation and ARE reporter activation (~10-fold compared to ≥100-fold activation). At similar H2O2 concentrations, both NRF2 protein synthesis and global protein synthesis were inhibited. The manganese porphyrin antioxidant MnTMPyP rescued both global protein synthesis and NRF2 protein synthesis from H2O2 inhibition and increased ARE reporter activation. Similar results were observed for the diphenol di-tert-butylhydroquinone (dtBHQ). In conclusion, induction of the NRF2/ARE pathway by H2O2 and dtBHQ-derived oxidative species can be limited by inhibition of NRF2 protein synthesis, likely by arrest of global protein synthesis.

2-Deoxy-d-ribose induces ferroptosis in renal tubular epithelial cells via ubiquitin-proteasome system-mediated xCT protein degradation

Ferroptosis is a novel form of cell death triggered by iron-dependent lipid peroxidation. Recent findings suggest that inhibiting system χc-induces ferroptosis by reducing intracellular cystine levels, and that ferroptosis in renal tubular epithelial cells (RTECs) contributes to acute kidney injury (AKI) and diabetic nephropathy. Moreover, 2-deoxy-d-ribose (dRib) has been shown to inhibit cystine uptake through xCT, the functional unit of system χc-, in β-cells. This study aimed to investigate if dRib induces ferroptosis in RTECs and identify the underlying mechanisms.dRib treatment reduced cystine uptake and glutathione (GSH) content, and increased intracellular levels of malondialdehyde (MDA), 4-hydroxynonenal (4-HNE), lipid reactive oxygen species (ROS), and cell death in both NRK-52E cells and primary cultured RTECs. However, treatment with inhibitors of ferroptosis, such as deferoxamine (DFO), ferrostatin-1 (Fer-1), and liproxstatin-1 (Lip-1), counteracted the effects of dRib on GSH, MDA, 4-HNE, and lipid ROS levels, as well as cell death. Additionally, 2-mercaptoethanol (2-ME) treatment or xCT gene overexpression protected against dRib-induced changes. Moreover, transmission electron microscopy revealed dRib-induced mitochondrial shrinkage, decrease in cristae number, and outer membrane rupture. Furthermore, dRib treatment upregulated the expression of genes associated with ferroptosis, and downregulated xCT protein expression. The decrease in xCT protein caused by dRib was consistently observed even when treated with the protein synthesis inhibitor cycloheximide. However, treatment with the proteasome inhibitor MG132 reversed the dRib-induced decrease in xCT protein expression. Additionally, dRib increased xCT protein ubiquitination.Overall, dRib induces ferroptosis in RTECs by degrading xCT protein through ubiquitin-proteasome system (UPS), resulting in reduced intracellular cystine uptake. Therefore, targeting the regulation of system χc-through UPS could be a potential therapeutic approach for AKI and diabetic nephropathy.

Antibacterial spectrum of medicinal plants: A review

Medicinal plants exerted their antibacterial effects through many mechanisms included inhibition of cell wall synthesis, inhibition of cell membrane synthesis, disruption of cytoplasmic membrane, inhibition of bacterial proteins synthesis, inhibition of nucleic acid synthesis, inhibition of energy metabolism, as well as inhibition of bacterial virulence factors. The current review highlighted the medicinal plants with antibacterial activity with their spectrum of action.

Fear response of rat pups to a non-aversive social stimulus: Evidence for the involvement of memory processes.

Learning processes in rats during early development are importantly mediated by the mother, which represents the primary source of environmental information. This study aimed to determine whether aversive early experiences can induce the expression of pups' fear responses toward a non-aversive stimulus as a consequence of a memory process. First, we determined pups' fear responses toward an anesthetized female after being exposed to this stimulus or an empty cage together with their mothers from Postnatal Day (PNDs) 1 to 4. Second, we evaluated if the administration of the protein synthesis inhibitor cycloheximide (CHX; 0.2mg/kg, subcutaneously (sc).) disrupted the reconsolidation processes and abolished the fear response on PND 9. Only female pups previously exposed to the female intruder expressed fear responses toward an anesthetized female on PND 8. CHX administration to female pups immediately after exposure to an anesthetized female on PND 8 suppressed fear responses on PND 9, indicating that the fear expression was the result of a memory process, probably mediated by the mother. These findings demonstrated that early experiences can shape responses to social stimuli in a sex-dependent manner and emphasize the critical role of the mother in influencing fear learning in a social context.

Targeted inhibition of protein synthesis renders cancer cells vulnerable to apoptosis by unfolded protein response

Cellular stress responses including the unfolded protein response (UPR) decide over the fate of an individual cell to ensure survival of the entire organism. During physiologic UPR counter-regulation, protective proteins are upregulated to prevent cell death. A similar strategy induces resistance to UPR in cancer. Therefore, we hypothesized that blocking protein synthesis following induction of UPR substantially enhances drug-induced apoptosis of malignant cells. In line, upregulation of the chaperone BiP was prevented by simultaneous arrest of protein synthesis in B cell malignancies. Cytotoxicity by immunotoxins—approved inhibitors of protein synthesis—was synergistically enhanced in combination with UPR-inducers in seven distinct hematologic and three solid tumor entities in vitro. Synergistic cell death depended on mitochondrial outer membrane permeabilization via BAK/BAX, which correlated with synergistic, IRE1α-dependent reduction of BID, accompanied by an additive fall of MCL-1. The strong synergy was reproduced in vivo against xenograft mouse models of mantle cell lymphoma, Burkitt’s lymphoma, and patient-derived acute lymphoblastic leukemia. In contrast, synergy was absent in blood cells of healthy donors suggesting a tumor-specific vulnerability. Together, these data support clinical evaluation of blocking stress response counter-regulation using inhibitors of protein synthesis as a novel therapeutic strategy.

Underfeeding Alters Brain Tissue Synthesis Rate in a Rat Brain Injury Model.

Brain injuries (BI) are highly disruptive, often having long lasting effects. Inadequate standard of care (SOC) energy support in the hospital leads to dietary energy deficiencies in BI patients. However, it is unclear how underfeeding (UF) affects protein synthesis post-BI. Therefore, in a rat model, we addressed the issue of UF on the protein fractional synthesis rate (fSR) post-BI. Compared to ad libitum (AL)-fed animals, we found that UF decreased protein synthesis in hind-limb skeletal muscle and cortical mitochondrial and structural proteins (p ≤ 0.05). BI significantly increased protein synthesis in the left and right cortices (p ≤ 0.05), but suppressed protein synthesis in the cerebellum (p ≤ 0.05) as compared to non-injured sham animals. Compared to underfeeding alone, UF in conjunction with BI (UF+BI) caused increased protein synthesis rates in mitochondrial, cytosolic, and whole-tissue proteins of the cortical brain regions. The increased rates of protein synthesis found in the UF+BI group were mitigated by AL feeding, demonstrating that caloric adequacy alleviates the effects of BI on protein dynamics in cortical and cerebellar brain regions. This research provides evidence that underfeeding has a negative impact on brain healing post-BI and that protein reserves in uninjured tissues are mobilized to support cortical tissue repair following BI.