No energy, no autophagy-Mechanisms and therapeutic implications of autophagic response energy requirements.

Autophagy is a lysosome-mediated self-degradation process of central importance for cellular quality control. It also provides macromolecule building blocks and substrates for energy metabolism during nutrient or energy deficiency, which are the main stimuli for autophagy induction. However, like most biological processes, autophagy itself requires ATP, and there is an energy threshold for its initiation and execution. We here present the first comprehensive review of this often-overlooked aspect of autophagy research. The studies in which ATP deficiency suppressed autophagy in vitro and in vivo were classified according to the energy pathway involved (oxidative phosphorylation or glycolysis). A mechanistic insight was provided by pinpointing the critical ATP-consuming autophagic events, including transcription/translation/interaction of autophagy-related molecules, autophagosome formation/elongation, autophagosome fusion with the lysosome, and lysosome acidification. The significance of energy-dependent fine-tuning of autophagic response for preserving the cell homeostasis, and potential implications for the therapy of cancer, autoimmunity, metabolic disorders, and neurodegeneration are discussed.

The role of Listeria monocytogenes PstA in ß-lactam resistance requires the cytochrome bd oxidase activity.

c-di-AMP is an essential second messenger that binds and regulates several proteins of different functions within bacterial cells. Among those, PstA is a structurally conserved c-di-AMP-binding protein, but its function is largely unknown. PstA is structurally similar to PII signal transduction proteins, although it specifically binds c-di-AMP rather than other PII ligands such as ATP and α-ketoglutarate. In Listeria monocytogenes, we found that PstA increases ß-lactam susceptibility at normal and low c-di-AMP levels, but increases ß-lactam resistance upon c-di-AMP accumulation. Examining a PstA mutant defective for c-di-AMP binding, we found the apo form of PstA to be toxic for ß-lactam resistance, and the c-di-AMP-bound form to be beneficial. Intriguingly, a role for PstA in ß-lactam resistance is only prominent in aerobic cultures, and largely diminished under hypoxic conditions, suggesting that PstA function is linked to aerobic metabolism. However, PstA does not control aerobic growth rate, and has a modest influence on the tricarboxylic acid cycle and membrane potential-an indicator of cellular respiration. The regulatory role of PstA in ß-lactam resistance is unrelated to reactive oxygen species or oxidative stress. Interestingly, during aerobic growth, PstA function requires the cytochrome bd oxidase (CydAB), a component of the respiratory electron transport chain. The requirement for CydAB might be related to its function in maintaining a membrane potential, or redox stress response activities. Altogether, we propose a model in which apo-PstA diminishes ß-lactam resistance by interacting with an effector protein, and this activity can be countered by c-di-AMP binding or a by-product of redox stress. IMPORTANCE: PstA is a structurally conserved c-di-AMP-binding protein that is broadly present among Firmicutes bacteria. Furthermore, PstA binds c-di-AMP at high affinity and specificity, indicating an important role in the c-di-AMP signaling network. However, the molecular function of PstA remains elusive. Our findings reveal contrasting roles of PstA in ß-lactam resistance depending on c-di-AMP-binding status. We also define physiological conditions for PstA function during aerobic growth. Future efforts can exploit these conditions to identify PstA interaction partners under ß-lactam stress.

The effect of intermittent fasting on preventing obesity-related early aging from a molecular and cellular perspective.

Obesity is a global health concern owing to its association with numerous degenerative diseases and the fact that it may lead to early aging. Various markers of aging, including telomere attrition, epigenetic alterations, altered protein homeostasis, mitochondrial dysfunction, cellular senescence, stem cell disorders, and intercellular communication, are influenced by obesity. Consequently, there is a critical need for safe and effective approaches to prevent obesity and mitigate the onset of premature aging. In recent years, intermittent fasting (IF), a dietary strategy that alternates between periods of fasting and feeding, has emerged as a promising dietary strategy that holds potential in counteracting the aging process associated with obesity. This article explores the molecular and cellular mechanisms through which IF affects obesity-related early aging. IF regulates various physiological processes and organ systems, including the liver, brain, muscles, intestines, blood, adipose tissues, endocrine system, and cardiovascular system. Moreover, IF modulates key signaling pathways such as AMP-activated protein kinase (AMPK), sirtuins, phosphatidylinositol 3-kinase (PI3K)/Akt, mammalian target of rapamycin (mTOR), and fork head box O (FOXO). By targeting these pathways, IF has the potential to attenuate aging phenotypes associated with obesity-related early aging. Overall, IF offers promising avenues for promoting healthier lifestyles and mitigating the premature aging process in individuals affected by obesity.

A MRSA mystery: how PBP4 and cyclic-di-AMP join forces against ß-lactam antibiotics.

The high-level resistance to next-generation ß-lactams frequently found in Staphylococcus aureus isolates lacking mec, which encodes the transpeptidase PBP2a traditionally associated with methicillin-resistant Staphylococcus aureus (MRSA), has remained incompletely understood for decades. A new study by Lai et al. found that the co-occurrence of mutations in pbp4 and gdpP, which respectively cause increased PBP4-mediated cell wall crosslinking and elevated cyclic-di-AMP levels, produces synergistic ß-lactam resistance rivaling that of PBP2a-producing MRSA (L.-Y. Lai, N. Satishkumar, S. Cardozo, V. Hemmadi, et al., mBio 15:e02889-23. 2024, https://doi.org/10.1128/mbio.02889-23). The combined mutations are sufficient to explain the high-level ß-lactam resistance of some mec-lacking strains, but the mechanism of synergy remains elusive and an avenue for further research. Importantly, the authors establish that co-occurrence of these mutations leads to antibiotic therapy failure in a Caenorhabditis elegans infection model. These results underscore the need to consider this unique and novel ß-lactam resistance mechanism during the clinical diagnosis of MRSA, rather than relying on mec as a diagnostic.

The involvement of CaMKKI in activating AMPKα in yesso scallop Patinopecten yessoensis under high temperature stress.

Calcium/calmodulin dependent protein kinase kinase (CaMKK), a highly conserved protein kinase, is involved in the downstream processes of various biological activities by phosphorylating and activating 5'-AMP-activated protein kinase (AMPK) in response to the increase of cytosolic-free calcium (Ca2+). In the present study, a CaMKKI was identified from Yesso scallop Patinopecten yessoensis. Its mRNA was ubiquitously expressed in haemocytes and all tested tissues with the highest expression level in mantle. The expression level of PyCaMKKI mRNA in adductor muscle was significantly upregulated at 1, 3 and 6 h after high temperature treatment (25 °C), which was 3.43-fold (p < 0.05), 5.25-fold (p < 0.05), and 5.70-fold (p < 0.05) of that in blank group, respectively. At 3 h after high temperature treatment (25 °C), the protein level of PyAMPKα, as well as the phosphorylation level of PyAMPKα at Thr170 in adductor muscle, and the positive co-localized fluorescence signals of PyCaMKKI and PyAMPKα in haemocyte all increased significantly (p < 0.05) compared to blank group (18 °C). The pull-down assay showed that rPyCaMKKI and rPyAMPKα could bind each other in vitro. After PyCaMKKI was silenced by siRNA, the mRNA and protein levels of PyCaMKKI and PyAMPKα, and the phosphorylation level of PyAMPKα at Thr170 in adductor muscle were significantly down-regulated (p < 0.05) compared with the negative control group receiving an injection of siRNA-NC. These results collectively suggested that PyCaMKKI was involved in the activation of PyAMPKα in response to high temperature stress and would be helpful for understanding the function of PyCaMKKI-PyAMPKα pathway in maintaining energy homeostasis under high temperature stress in scallops.

SIRT3/AMPK signaling pathway regulates lipid metabolism and improves vulnerability to atrial fibrillation in Dahl salt sensitive rats.

BACKGROUND: Hypertension may result in atrial fibrillation (AF) and lipid metabolism disorders. The Sirtuins3 (SIRT3) / AMP-activated protein kinase (AMPK) signaling pathway has the capacity to regulate lipid metabolism disorders and the onset of AF. We hypothesize that the SIRT3/AMPK signaling pathway suppresses lipid metabolism disorders, thereby mitigating salt-sensitive hypertension (SSHT)-induced susceptibility to AF. METHODS: The study involved 7-week-old male Dahl salt-sensitive that were fed either high-salt diet (8% NaCl; DSH group) or normal diet (0.3% NaCl; DSN group). Then DSH group were administered either oral metformin (MET, an AMPK agonist) or intraperitoneal injection of Honokiol (HK, a SIRT3 agonist). This experimental model allowed for the measurement of SBP, the expression levels of lipid metabolism-related biomarker, pathological examination of atrial fibrosis and lipid accumulation, as well as AF inducibility and AF duration. RESULTS: DSH decrease SIRT3, phosphorylation-AMPK and VLCAD expression, increased FASN and FABP4 expression and concentrations of FFA and TG, atrial fibrosis and lipid accumulation in atrial tissue, enhanced level of SBP, promoted AF induction rate and prolonged AF duration, which are blocked by MET and HK. Our results also showed that the degree of atrial fibrosis was negatively correlated with VLCAD expression, but positively correlated with the expression of FASN and FABP4. CONCLUSIONS: We have confirmed that high-salt diet can result in hypertension, associated atrial tissue lipid metabolism dysfunction. This condition is linked to the inhibition of the SIRT3/AMPK signaling pathway, which plays a significant role in the progression of susceptibility to AF in SSHT rats.

Assessment of the evidence yield for the calibrated PP3/BP4 computational recommendations.

PURPOSE: To investigate the number of rare missense variants observed in human genome sequences by ACMG/AMP PP3/BP4 evidence strength, following the calibrated PP3/BP4 computational recommendations. METHODS: Missense variants from the genome sequences of 300 probands from the Rare Genomes Project with suspected rare disease were analyzed using computational prediction tools able to reach PP3_Strong and BP4_Moderate evidence strengths (BayesDel, MutPred2, REVEL, and VEST4). The numbers of variants at each evidence strength were analyzed across disease-associated genes and genome-wide. RESULTS: From a median of 75.5 rare (≤1% allele frequency) missense variants in disease-associated genes per proband, a median of one reached PP3_Strong, 3-5 PP3_Moderate, and 3-5 PP3_Supporting. Most were allocated BP4 evidence (median 41-49 per proband) or were indeterminate (median 17.5-19 per proband). Extending the analysis to all protein-coding genes genome-wide, the number of PP3_Strong variants increased approximately 2.6-fold compared to disease-associated genes, with a median per proband of 1-3 PP3_Strong, 8-16 PP3_Moderate, and 10-17 PP3_Supporting. CONCLUSION: A small number of variants per proband reached PP3_Strong and PP3_Moderate in 3,424 disease-associated genes, and though not the intended use of the recommendations, also genome-wide. Use of PP3/BP4 evidence as recommended from calibrated computational prediction tools in the clinical diagnostic laboratory is unlikely to inappropriately contribute to the classification of an excessive number of variants as Pathogenic or Likely Pathogenic by ACMG/AMP rules.

Activation AMPK in Hypothalamic Paraventricular Nucleus Improves Renovascular Hypertension Through ERK1/2-NF-κB Pathway.

Hypertension is a globally prevalent disease, but the pathogenesis remains largely unclear. AMP-activated protein kinase (AMPK) is a nutrition-sensitive signal of cellular energy metabolism, which has a certain influence on the development of hypertension. Previously, we found a down-regulation of the phosphorylated (p-) form of AMPK, and the up-regulation of the angiotensin II type 1 receptor (AT1-R) and that of p-ERK1/2 in the hypothalamic paraventricular nucleus (PVN) of hypertensive rats. However, the exact mechanism underlying the relationship between AMPK and AT1-R in the PVN during hypertension remains unclear. Thus, we hypothesized that AMPK modulates AT1-R through the ERK1/2-NF-κB pathway in the PVN, thereby inhibiting sympathetic nerve activity and improving hypertension. To examine this hypothesis, we employed a renovascular hypertensive animal model developed via two-kidney, one-clip (2K1C) and sham-operated (SHAM). Artificial cerebrospinal fluid (aCSF), used as vehicle, or 5-amino-1-ß-D-ribofuranosyl-imidazole-4-carboxamide (AICAR, an AMPK activator, 60 µg/day) was microinjected bilaterally in the PVN of these rats for 4 weeks. In 2K1C rats, there an increase in systolic blood pressure (SBP) and circulating norepinephrine (NE). Also, the hypertensive rats had lowered expression of p-AMPK and p-AMPK/AMPK, elevated expression of p-ERK1/2, p-ERK1/2/ERK1/2 and AT1-R, increased NF-κB p65 activity in the PVN compared with the levels of these biomarkers in SHAM rats. Four weeks of bilateral PVN injection of AMPK activator AICAR, attenuated the NE level and SBP, increased the expression of p-AMPK and p-AMPK/AMPK, lessened the NF-κB p65 activity, decreased the expression of p-ERK1/2, p-ERK1/2/ERK1/2 and AT1-R in the PVN of 2K1C rats. Data from this study imply that the activation of AMPK within the PVN suppressed AT1-R expression through inhibiting the ERK1/2-NF-κB pathway, decreased the activity of the sympathetic nervous system, improved hypertension.

A network of acetyl phosphate-dependent modification modulates c-di-AMP homeostasis in Actinobacteria.

Cyclic purine nucleotides are important signal transduction molecules across all domains of life. 3',5'-cyclic di-adenosine monophosphate (c-di-AMP) has roles in both prokaryotes and eukaryotes, while the signals that adjust intracellular c-di-AMP and the molecular machinery enabling a network-wide homeostatic response remain largely unknown. Here, we present evidence for an acetyl phosphate (AcP)-governed network responsible for c-di-AMP homeostasis through two distinct substrates, the diadenylate cyclase DNA integrity scanning protein (DisA) and its newly identified transcriptional repressor, DasR. Correspondingly, we found that AcP-induced acetylation exerts these regulatory actions by disrupting protein multimerization, thus impairing c-di-AMP synthesis via K66 acetylation of DisA. Conversely, the transcriptional inhibition of disA was relieved during DasR acetylation at K78. These findings establish a pivotal physiological role for AcP as a mediator to balance c-di-AMP homeostasis. Further studies revealed that acetylated DisA and DasR undergo conformational changes that play crucial roles in differentiation. Considering the broad distribution of AcP-induced acetylation in response to environmental stress, as well as the high conservation of the identified key sites, we propose that this unique regulation of c-di-AMP homeostasis may constitute a fundamental property of central circuits in Actinobacteria and thus the global control of cellular physiology.IMPORTANCESince the identification of c-di-AMP is required for bacterial growth and cellular physiology, a major challenge is the cell signals and stimuli that feed into the decision-making process of c-di-AMP concentration and how that information is integrated into the regulatory pathways. Using the bacterium Saccharopolyspora erythraea as a model, we established that AcP-dependent acetylation of the diadenylate cyclase DisA and its newly identified transcriptional repressor DasR is involved in coordinating environmental and intracellular signals, which are crucial for c-di-AMP homeostasis. Specifically, DisA acetylated at K66 directly inactivates its diadenylate cyclase activity, hence the production of c-di-AMP, whereas DasR acetylation at K78 leads to increased disA expression and c-di-AMP levels. Thus, AcP represents an essential molecular switch in c-di-AMP maintenance, responding to environmental changes and possibly hampering efficient development. Therefore, AcP-mediated posttranslational processes constitute a network beyond the usual and well-characterized synthetase/hydrolase governing c-di-AMP homeostasis.

Docosahexaenoic acid supplementation during porcine oocyte in vitro maturation improves oocyte quality and embryonic development by enhancing the homeostasis of energy metabolism.

Although supplementation with docosahexaenoic acid (DHA) during porcine oocyte IVM is well-established, the available data are limited due to the lack of consistency. Moreover, to our knowledge, the anti-oxidant effects of DHA on porcine oocytes have not been reported. Hence, this study aimed to examine the effects of DHA supplementation on the regulation of energy metabolism during porcine oocyte maturation to improve oocyte maturation and embryonic development. By supplementing the IVM medium with various DHA concentrations, 25 µM DHA was identified as the optimal concentration which improved intraoocyte glutathione content and enhanced embryonic development after parthenogenesis. Compared to embryos derived from the control group, those derived from SCNT or IVF showed significantly improved blastocyst formation upon DHA supplementation during IVM. In addition, various transcription factors associated with oocyte development and apoptosis in mature oocytes were beneficially regulated in the DHA-treated oocytes. Moreover, DHA improved the AMP-activated protein kinase (AMPK)-regulatory ability of porcine oocytes and ameliorated nuclear maturation and embryonic development, which were decreased by artificially downregulating AMPK. To our knowledge, this is the first study to examine the effects of DHA as an AMPK regulator on oocyte maturation and embryo development in pigs. Furthermore, DHA addition to the IVM medium upregulated the relative expression of genes associated with mitochondrial potential and lipid metabolism. Therefore, the membrane potential of mitochondria (evaluated based on the JC-1 aggregate/JC-1 monomer ratio) and the levels of fatty acids and lipid droplets in matured oocytes increased, resulting in increased ATP synthesis. In conclusion, the DHA treatment of porcine oocytes with 25 µM DHA during IVM enhances the homeostasis of energy metabolism by improving mitochondrial function and lipid metabolism, leading to improved quality of matured oocytes and enhanced embryonic developmental potential of in vitro produced (IVP) embryos. Thus, 25 µM DHA supplementation could serve as a tool for improving the quality of IVP embryos. The study findings provide a basis for further research on improving the production efficiency of cloned animals by securing high-quality matured oocytes and enhancing energy metabolism in mammalian oocytes, including those of pigs.

Oral Preexposure Prophylaxis Uptake and Discontinuation in the HIV Vaccine Trials Network 704/HIV Prevention Trials Network 085 Study: Implications for Biomedical Human Immunodeficiency Virus Prevention Trials.

Background: HIV Vaccine Trials Network (HVTN) 704/085, a placebo-controlled clinical trial assessing the efficacy of VRC01 broadly neutralizing antibody infusion for HIV prevention, offered oral preexposure prophylaxis (PrEP) as the standard of prevention at no cost to participants. Methods: We characterized features of- identified factors associated with- PrEP initiation and discontinuation, and the effects of PrEP initiation on HIV incidence. Results: Of 2221 participants, 31.8% initiated oral PrEP during study follow-up, with the highest proportion of PrEP initiations in Brazil (83.2%) and the United States (US) (54.2%). Prior PrEP use was associated with PrEP initiation (hazard ratio [HR], 2.22 [95% confidence interval {CI}, 1.25-3.95]). Participants from Switzerland (HR, 0.5 [95% CI, .3-1.0]) and Peru (HR, 0.08 [95% CI, .06-.1]) had lower likelihood of PrEP initiation compared to the US, while participants from Brazil had higher likelihood (HR, 2.6 [95% CI, 2.0-3.3]). In the US, PrEP initiation was lower in areas with higher unmet need for PrEP (HR, 0.9 per 5 units [95% CI, 0.8-1.0]). PrEP initiators had 58% less risk of acquiring HIV than PrEP noninitiators. Among PrEP initiators, 34.4% discontinued PrEP during study follow-up. Brazil had 63% less likelihood of PrEP discontinuation than the US (HR, 0.37 [95% CI, .22-.60]). Conclusions: When included as standard of prevention in HVTN 704/085, oral PrEP utilization patterns mirrored those observed in real-life settings. Variable effects of oral PrEP on HIV outcomes in clinical trials may be expected based on regional differences in oral PrEP use.

Heterologous expression of the insect SVWC peptide WHIS1 inhibits Candida albicans invasion into A549 and HeLa epithelial cells.

Candida albicans (C. albicans), a microbe commonly isolated from Candida vaginitis patients with vaginal tract infections, transforms from yeast to hyphae and produces many toxins, adhesins, and invasins, as well as C. albicans biofilms resistant to antifungal antibiotic treatment. Effective agents against this pathogen are urgently needed. Antimicrobial peptides (AMPs) have been used to cure inflammation and infectious diseases. In this study, we isolated whole housefly larvae insect SVWC peptide 1 (WHIS1), a novel insect single von Willebrand factor C-domain protein (SVWC) peptide from whole housefly larvae. The expression pattern of WHIS1 showed a response to the stimulation of C. albicans. In contrast to other SVWC members, which function as antiviral peptides, interferon (IFN) analogs or pathogen recognition receptors (PRRs), which are the prokaryotically expressed MdWHIS1 protein, inhibit the growth of C. albicans. Eukaryotic heterologous expression of WHIS1 inhibited C. albicans invasion into A549 and HeLa cells. The heterologous expression of WHIS1 clearly inhibited hyphal formation both extracellularly and intracellularly. Furthermore, the mechanism of WHIS1 has demonstrated that it downregulates all key hyphal formation factors (ALS1, ALS3, ALS5, ECE1, HWP1, HGC1, EFG1, and ZAP1) both extracellularly and intracellularly. These data showed that heterologously expressed WHIS1 inhibits C. albicans invasion into epithelial cells by affecting hyphal formation and adhesion factor-related gene expression. These findings provide new potential drug candidates for treating C. albicans infection.

Inhibitory Action of 1,3,5-Trihydroxybenzene on UVB-Induced NADPH Oxidase 4 through AMPK and JNK Signaling Pathways.

Specific sensitivity of the skin to ultraviolet B (UVB) rays is one of the mechanisms responsible for widespread skin damage. This study tested whether 1,3,5-trihydroxybenzene (THB), a compound abundant in marine products, might inhibit UVB radiation-induced NADPH oxidase 4 (NOX4) in both human HaCaT keratinocytes and mouse dorsal skin and explore its cytoprotective mechanism. The mechanism of action was determined using western blotting, immunocytochemistry, NADP+/NADPH assay, reactive oxygen species (ROS) detection, and cell viability assay. THB attenuated UVB-induced NOX4 expression both in vitro and in vivo, and suppressed UVB-induced ROS generation via NADP+ production, resulting in increased cell viability with decreased apoptosis. THB also reduced the expression of UVB-induced phosphorylated AMP-activated protein kinase (AMPK) and phosphorylated c-Jun N-terminal kinase (JNK). THB suppressed UVB-induced NOX4 expression and ROS generation by inhibiting AMPK and JNK signaling pathways, thereby inhibiting cellular damage. These results showed that THB could be developed as a UV protectant.

Unlocking roles of cationic and aromatic residues in peptide amphiphiles in treating drug-resistant gram-positive pathogens.

Multidrug resistance (MDR) is a rising threat to global health because the number of essential antibiotics used for treating MDR infections is increasingly compromised. In this work we report a group of new amphiphilic peptides (AMPs) derived from the well-studied G3 (G(IIKK)3I-NH2) to fight infections from Gram-positive bacteria including susceptible Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA), focusing on membrane interactions. Time-dependent killing experiments revealed that substitutions of II by WW (GWK), II by FF (GFK) and KK by RR (GIR) resulted in improved bactericidal efficiencies compared to G3 (GIK) on both S. aureus and MRSA, with the order of GWK > GIR > GFK > GIK. Electronic microscopy imaging revealed structural disruptions of AMP binding to bacterial cell walls. Fluorescence assays including AMP binding to anionic lipoteichoic acids (LTA) in cell-free and cell systems indicated concentration and time-dependent membrane destabilization associated with bacterial killing. Furthermore, AMP's binding to anionic plasma membrane via similar fluorescence assays revealed a different extent of membrane depolarization and leakage. These observations were supported by the penetration of AMPs into the LTA barrier and the subsequent structural compromise to the cytoplasmic membrane as revealed from SANS (small angle neutron scattering). Both experiments and molecular dynamics (MD) simulations revealed that GWK and GIR could make the membrane more rigid but less effective in diffusive efficiency than GIK and GFK through forming intramembrane peptide nanoaggregates associated with hydrophobic mismatch and formation of fluidic and rigid patches. The reported peptide-aggregate-induced phase-separation emerged as a crucial factor in accelerated membrane disintegration and fast bacterial killing. This work has demonstrated the importance of membrane interactions to the development of more effective AMPs and the relevance of the approaches as reported in assisting this area of research.

[Retracted] Effect of metformin on cell proliferation, apoptosis, migration and invasion in A172 glioma cells and its mechanisms.

Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that the cell invasion and migration assay data shown in Fig. 6 and the cell proliferation assay experiments shown in Fig. 2 were strikingly similar to data appearing in different form in other articles by different authors; furthermore, in Fig. 2, for the '10 mM metformin' experiment, certain of the glioma cells appeared to be strikingly similar to other cells contained within the same data panels. Owing to the fact that the contentious data in the above article had already been published elsewhere or were under consideration for publication prior to its submission to Molecular Medicine Reports, and owing to concerns with the authenticity of certain of the data, the Editor has decided that this paper should be retracted from the Journal. The authors were asked for an explanation to account for these concerns, but the Editorial Office did not receive a reply. The Editor apologizes to the readership for any inconvenience caused. [Molecular Medicine Reports 20: 887­894, 2019; DOI: 10.3892/mmr.2019.10369].

A review on the diversity of antimicrobial peptides and genome mining strategies for their prediction.

Antibiotic resistance has become one of the most serious threats to human health in recent years. In response to the increasing microbial resistance to the antibiotics currently available, it is imperative to develop new antibiotics or explore new approaches to combat antibiotic resistance. Antimicrobial peptides (AMPs) have shown considerable promise in this regard, as the microbes develop low or no resistance against them. The discovery and development of AMPs still confront numerous obstacles such as finding a target, developing assays, and identifying hits and leads, which are time-consuming processes, making it difficult to reach the market. However, with the advent of genome mining, new antibiotics could be discovered efficiently using tools such as BAGEL, antiSMASH, RODEO, etc., providing hope for better treatment of diseases in the future. Computational methods used in genome mining automatically detect and annotate biosynthetic gene clusters in genomic data, making it a useful tool in natural product discovery. This review aims to shed light on the history, diversity, and mechanisms of action of AMPs and the data on new AMPs identified by traditional as well as genome mining strategies. It further substantiates the various phases of clinical trials for some AMPs, as well as an overview of genome mining databases and tools built expressly for AMP discovery. In light of the recent advancements, it is evident that targeted genome mining stands as a beacon of hope, offering immense potential to expedite the discovery of novel antimicrobials.

Bacterial cell volume regulation and the importance of cyclic di-AMP.

SUMMARYNucleotide-derived second messengers are present in all domains of life. In prokaryotes, most of their functionality is associated with general lifestyle and metabolic adaptations, often in response to environmental fluctuations of physical parameters. In the last two decades, cyclic di-AMP has emerged as an important signaling nucleotide in many prokaryotic lineages, including Firmicutes, Actinobacteria, and Cyanobacteria. Its importance is highlighted by the fact that both the lack and overproduction of cyclic di-AMP affect viability of prokaryotes that utilize cyclic di-AMP, and that it generates a strong innate immune response in eukaryotes. In bacteria that produce the second messenger, most molecular targets of cyclic di-AMP are associated with cell volume control. Besides, other evidence links the second messenger to cell wall remodeling, DNA damage repair, sporulation, central metabolism, and the regulation of glycogen turnover. In this review, we take a biochemical, quantitative approach to address the main cellular processes that are directly regulated by cyclic di-AMP and show that these processes are very connected and require regulation of a similar set of proteins to which cyclic di-AMP binds. Altogether, we argue that cyclic di-AMP is a master regulator of cell volume and that other cellular processes can be connected with cyclic di-AMP through this core function. We further highlight important directions in which the cyclic di-AMP field has to develop to gain a full understanding of the cyclic di-AMP signaling network and why some processes are regulated, while others are not.

Environmentally responsive hydrogel promotes vascular normalization to enhance STING anti-tumor immunity.

The immunosuppressive microenvironment of malignant tumors severely hampers the effectiveness of anti-tumor therapy. Moreover, abnormal tumor vasculature interacts with immune cells, forming a vicious cycle that further interferes with anti-tumor immunity and promotes tumor progression. Our pre-basic found excellent anti-tumor effects of c-di-AMP and RRx-001, respectively, and we further explored whether they could be combined synergistically for anti-tumor immunotherapy. We chose to load these two drugs on PVA-TSPBA hydrogel scaffolds that expressly release drugs within the tumor microenvironment by in situ injection. Studies have shown that c-di-AMP activates the STING pathway, enhances immune cell infiltration, and reverses tumor immunosuppression. Meanwhile, RRx-001 releases nitric oxide, which increases oxidative stress injury in tumor cells and promotes apoptosis. Moreover, the combination of the two presented more powerful pro-vascular normalization and reversed tumor immunosuppression than the drug alone. This study demonstrates a new design option for anti-tumor combination therapy and the potential of tumor environmentally responsive hydrogel scaffolds in combination with anti-tumor immunotherapy.