Synthesis of histidine, serine and folic acid-functionalized and boron and iron-doped graphene quantum dot with excellent optical behavior and peroxidase-like activity for colorimetric and fluorescence detection of H2O2 in food.

Low fluorescence under visible light excitation and catalytic activity limit many applications of graphene quantum dots in optical detection, biosensing, catalysis and biomedical. The paper reports design and synthesis of histidine, serine and folic acid-functionalized and boron and iron-doped graphene quantum dot (Fe/B-GQD-HSF). The Fe/B-GQD-HSF shows excellent fluorescence behavior and peroxidase-like activity. Excitation of 330 nm ultraviolet light produces the strongest blue fluorescence and excitation of 480 nm visible light produces the strongest yellow fluorescence. The specific activity reaches 92.67 U g-1, which is higher than that of other graphene quantum dots. The Fe/B-GQD-HSF can catalyze oxidation of 3,3',5,5'-tetramethylbenzidine with H2O2 to form blue compound. Based on this, it was used for colorimetric and fluorescence detection of H2O2. The absorbance at 652 nm linearly increases with the increase of H2O2 concentration between 0.5 and 100 µM with detection limit of 0.43 µM. The fluorescence signal linearly decreases with the increase of H2O2 concentration between 0.05 and 100 µM with detection limit of 0.035 µM. The analytical method has been satisfactorily applied in detection of H2O2 in food. The study also paves one way for design and synthesis of functional graphene quantum dots with ideal fluorescence behavior and catalytic activity.

Orthogonal Serine Integrases Enable Scalable Gene Storage Cascades in Bacterial Genome.

Genome integration enables host organisms to stably carry heterologous DNA messages, introducing new genotypes and phenotypes for expanded applications. While several genome integration approaches have been reported, a scalable tool for DNA message storage within site-specific genome landing pads is still lacking. Here, we introduce an iterative genome integration method utilizing orthogonal serine integrases, enabling the stable storage of multiple heterologous genes in the chromosome of Escherichia coli MG1655. By leveraging serine integrases TP901-1, Bxb1, and PhiC31, along with engineered integration vectors, we demonstrate high-efficiency, marker-free integration of DNA fragments up to 13 kb in length. To further simplify the procedure, we then develop a streamlined integration method and showcase the system's versatility by constructing an engineered E. coli strain capable of storing and expressing multiple genes from diverse species. Additionally, we illustrate the potential utility of these engineered strains for synthetic biology applications, including in vivo and in vitro protein expression. Our work extends the application scope of serine integrases for scalable gene integration cascades, with implications for genome manipulation and gene storage applications in synthetic biology.

A metabolic crosstalk between liposarcoma and muscle sustains tumor growth.

Dedifferentiated and Well-differentiated liposarcoma are characterized by a systematic amplification of the Murine Double Minute 2 (MDM2) oncogene. We demonstrate that p53-independent metabolic functions of chromatin-bound MDM2 are exacerbated in liposarcoma and mediate an addiction to serine metabolism to sustain tumor growth. However, the origin of exogenous serine remains unclear. Here, we show that elevated serine levels in mice harboring liposarcoma-patient derived xenograft, released by distant muscle is essential for liposarcoma cell survival. Repressing interleukine-6 expression, or treating liposarcoma cells with Food and Drugs Administration (FDA) approved anti-interleukine-6 monoclonal antibody, decreases de novo serine synthesis in muscle, impairs proliferation, and increases cell death in vitro and in vivo. This work reveals a metabolic crosstalk between muscle and liposarcoma tumor and identifies anti-interleukine-6 as a plausible treatment for liposarcoma patients.

Thr4 phosphorylation on RNA Pol II occurs at early transcription regulating 3'-end processing.

RNA polymerase II relies on a repetitive sequence domain (YSPTSPS) within its largest subunit to orchestrate transcription. While phosphorylation on serine-2/serine-5 of the carboxyl-terminal heptad repeats is well established, threonine-4's role remains enigmatic. Paradoxically, threonine-4 phosphorylation was only detected after transcription end sites despite functionally implicated in pausing, elongation, termination, and messenger RNA processing. Our investigation revealed that threonine-4 phosphorylation detection was obstructed by flanking serine-5 phosphorylation at the onset of transcription, which can be removed selectively. Subsequent proteomic analyses identified many proteins recruited to transcription via threonine-4 phosphorylation, which previously were attributed to serine-2. Loss of threonine-4 phosphorylation greatly reduces serine-2 phosphorylation, revealing a cross-talk between the two marks. Last, the function analysis of the threonine-4 phosphorylation highlighted its role in alternative 3'-end processing within pro-proliferative genes. Our findings unveil the true genomic location of this evolutionarily conserved phosphorylation mark and prompt a reassessment of functional assignments of the carboxyl-terminal domain.

Tryptophan production by catalysis of a putative tryptophan synthase protein.

Essential amino acid, tryptophan which intake from food plays a critical role in numerous metabolic functions, exhibiting extensive biological functions and applications. Tryptophan is beneficial for the food sector by enhancing nutritional content and promoting the development of functional foods. A putative gene encoding tryptophan synthase was the first identified in Sphingobacterium soilsilvae Em02, a cellulosic bacterium making it inherently more environmentally friendly. The gene was cloned and expressed in exogenous host Escherichia coli, to elucidate its function. The recombinant tryptophan synthase with a molecular weight 42 KDa was expressed in soluble component. The enzymatic activity to tryptophan synthase in vivo was assessed using indole and L-serine and purified tryptophan synthase. The optimum enzymatic activity for tryptophan synthase was recorded at 50 ºC and pH 7.0, which was improved in the presence of metal ions Mg2+, Sr2+ and Mn2+, whereas Cu2+, Zn2+ and Co2+ proved to be inhibitory. Using site-directed mutagenesis, the consensus pattern HK-S-[GGGSN]-E-S in the tryptophan synthase was demonstrated with K100Q, S202A, G246A, E361A and S385A as the active sites. Tryptophan synthase has been demonstrated to possess the defining characteristics of the ß-subunits. The tryptophan synthase may eventually be useful for tryptophan production on a larger scale. Its diverse applications highlight the potential for improving both the quality and health benefits of food products, making it an essential component in advancing food science and technology.

Oncofetal TRIM71 drives liver cancer carcinogenesis through remodeling CEBPA-mediated serine/glycine metabolism.

Rationale: Tumor cells remodel transcriptome to construct an ecosystem with stemness features, which maintains tumor growth and highly malignant characteristics. However, the core regulatory factors involved in this process still need to be further discovered. Methods: Single cell RNA-sequncing (scRNA-seq) and bulk RNA-sequencing profiles derived from fetal liver, normal liver, liver tumors, and their adjacent samples were collected to analyze the ecosystem of liver cancer. Mouse models were established to identify molecular functions of oncofetal-related oncogenes using hydrodynamic tail vein injection. Results: We found that liver cancer rebuilt oncofetal ecosystem to maintain malignant features. Interestingly, we identified a group of RNA-binding proteins (RBPs) that were highly overexpressed with oncofetal features. Among them, TRIM71 was specifically expressed in liver cancers and was associated with poor outcomes. TRIM71 drove the carcinogenesis of hepatocellular carcinoma (HCC), and knockdown of TRIM71 significantly abolished liver cancer cell proliferation. Mechanistically, TRIM71 formed a protein complex with IGF2BP1, bound to and stabilized the mRNA of CEBPA in an m6A-dependent manner, enhance the serine/glycine metabolic pathway, and ultimately promoted liver cancer progression. Furthermore, we identified that all-trans-retinoic acid (ATRA) combined with e1A binding protein p300 (EP300) inhibitor A-485 repressed TRIM71, attenuated glycine/serine metabolism, and inhibited liver cancer cell proliferation with high TRIM71 levels. Conclusions: We demonstrated the oncofetal status in liver cancer and highlighted the crucial role of TRIM71 and provided potential therapeutic strategies and liver cancer-specific biomarker for liver cancer patients.

MerlinS13 phosphorylation regulates meningioma Wnt signaling and magnetic resonance imaging features.

Meningiomas are associated with inactivation of NF2/Merlin, but approximately one-third of meningiomas with favorable clinical outcomes retain Merlin expression. Biochemical mechanisms underlying Merlin-intact meningioma growth are incompletely understood, and non-invasive biomarkers that may be used to guide treatment de-escalation or imaging surveillance are lacking. Here, we use single-cell RNA sequencing, proximity-labeling proteomic mass spectrometry, mechanistic and functional approaches, and magnetic resonance imaging (MRI) across meningioma xenografts and patients to define biochemical mechanisms and an imaging biomarker that underlie Merlin-intact meningiomas. We find Merlin serine 13 (S13) dephosphorylation drives meningioma Wnt signaling and tumor growth by attenuating inhibitory interactions with ß-catenin and activating the Wnt pathway. MRI analyses show Merlin-intact meningiomas with S13 phosphorylation and favorable clinical outcomes are associated with high apparent diffusion coefficient (ADC). These results define mechanisms underlying a potential imaging biomarker that could be used to guide treatment de-escalation or imaging surveillance for patients with Merlin-intact meningiomas.

Structure and Intercalation of Cysteine-Asparagine-Serine Peptide into Montmorillonite as an Anti-Inflammatory Agent Preparation-A DFT Study.

Peptides are receiving significant attention in pharmaceutical sciences due to their applications as anti-inflammatory drugs; however, many aspects of their interactions and mechanisms at the molecular level are not well-known. This work explores the molecular structure of two peptides-(i) cysteine (Cys)-asparagine (Asn)-serine (Ser) (CNS) as a molecule in the gas phase and solvated in water in zwitterion form, and (ii) the crystal structure of the dipeptide serine-asparagine (SN), a reliable peptide indication whose experimental cell parameters are well known. A search was performed by means of atomistic calculations based on density functional theory (DFT). These calculations matched the experimental crystal structure of SN, validating the CNS results and useful for assignments of our experimental spectroscopic IR bands. Our calculations also explore the intercalation of CNS into the interlayer space of montmorillonite (MNT). Our quantum mechanical calculations show that the conformations of these peptides change significantly during intercalation into the confined interlayer space of MNT. This intercalation is energetically favorable, indicating that this process can be a useful preparation for therapeutic anti-inflammatory applications and showing high stability and controlled release processes.

A cocktail of histidine, carnosine, cysteine and serine reduces adiposity and improves metabolic health and adipose tissue immunometabolic function in ovariectomized rats.

Many women have sought alternative therapies to address menopause. Recently, a multi-ingredient supplement (MIS) containing L-histidine, L-carnosine, L-serine, and L-cysteine has been shown to be effective at ameliorating hepatic steatosis (HS) in ovariectomized (OVX) rats, a postmenopausal oestrogen deficiency model. Considering that HS frequently accompanies obesity, which often occurs during menopause, we aimed to investigate the effects of this MIS for 8 weeks in OVX rats. Twenty OVX rats were orally supplemented with either MIS (OVX-MIS) or vehicle (OVX). Ten OVX rats received vehicle orally along with subcutaneous injections of 17ß-oestradiol (OVX-E2), whereas 10 rats underwent a sham operation and received oral and injected vehicles (control group). MIS consumption partly counteracted the fat mass accretion observed in OVX animals, leading to decreased total fat mass, adiposity index and retroperitoneal white adipose tissue (RWAT) adipocyte hypertrophy. OVX-MIS rats also displayed increased lean mass and lean/fat ratio, suggesting a healthier body composition, similar to the results reported for OVX-E2 animals. MIS consumption decreased the circulating levels of the proinflammatory marker CRP, the total cholesterol-to-HDL-cholesterol ratio and the leptin-to-adiponectin ratio, a biomarker of diabetes risk and metabolic syndrome. RWAT transcriptomics indicated that MIS favourably regulated genes involved in adipocyte structure and morphology, cell fate determination and differentiation, glucose/insulin homeostasis, inflammation, response to stress and oxidative phosphorylation, which may be mechanisms underlying the beneficial effects described for OVX-MIS rats. Our results pave the way for using this MIS formulation to improve the body composition and immunometabolic health of menopausal women.

Effect of arginine, glycine + serine concentrations, and guanidinoacetic acid supplementation in vegetable-based diets for chickens.

The study investigated guanidinoacetic acid (GAA) supplementation with varying dietary digestible arginine (Arg) and glycine+serine (Gly+Ser) concentrations in the starter phase, exploring respective carry-over effects on growth performance, blood chemistry, incidence of pectoral myopathies and proximate composition in broilers. A total of 2,800 one-day-old male broiler chicks were distributed in a central composite design with 2 factors and double experimental mesh, represented by supplementation or omission of 0.6 g per kg of GAA, with a central point represented by 107% of Arg and 147% of Gly+Ser, 4 factorial points (combinations of Arg/Gly+Ser concentrations: 96.4/132.5%; 117.6/132.5%; 96.4/161.5%, and 117.6/132.5%), and 4 axial points (combinations of axial points estimated for Arg and Gly+Ser, with the central points of 92/147%; 122/147%; 107/126.5, and 107/167.5%), totaling 18 treatments, 4 repetitions to factorial and axial points, 24 replicates to the central point, and 25 birds per pen. Feed conversion ratio (FCR) from d 1 to 10 had a linear response (P = 0.009) for the decreasing Arg content and a quadratic response (P = 0.047) for Gly+Ser concentrations. Broilers supplemented GAA had lower FCR compared with nonsupplemented groups from d 1 to 10 (P = 0.048) and d 1 to 42 (P = 0.026). Aspartate aminotransferase (AST) exhibited increasing and decreasing linear effects as a function of Arg (P = 0.008) and Gly+Ser (P = 0.020) concentrations, respectively. Guanidinoacetic acid decreased serum AST (P = 0.028). Guanidinoacetic acid reduced moderate + severe (P = 0.039) and mild (P = 0.015) Wooden Breast scores. The occurrence of normal White Striping increased (P = 0.002), while severe score was reduced (P = 0.029) with GAA supplementation. In conclusion, increased digestible Arg:Lys and 14% and 6% above the recommendations (107% and 147%), respectively, provided improved FCR during the starter phase. Dietary GAA supplementation (0.6 g per kg) improved FCR, reduced severity of breast myopathies and appears to have reduced muscle damage in broilers fed plant-based diets.

The unique catalytic properties of PSAT1 mediate metabolic adaptation to glutamine blockade.

Cultured cancer cells frequently rely on the consumption of glutamine and its subsequent hydrolysis by glutaminase (GLS). However, this metabolic addiction can be lost in the tumour microenvironment, rendering GLS inhibitors ineffective in the clinic. Here we show that glutamine-addicted breast cancer cells adapt to chronic glutamine starvation, or GLS inhibition, via AMPK-mediated upregulation of the serine synthesis pathway (SSP). In this context, the key product of the SSP is not serine, but α-ketoglutarate (α-KG). Mechanistically, we find that phosphoserine aminotransferase 1 (PSAT1) has a unique capacity for sustained α-KG production when glutamate is depleted. Breast cancer cells with resistance to glutamine starvation or GLS inhibition are highly dependent on SSP-supplied α-KG. Accordingly, inhibition of the SSP prevents adaptation to glutamine blockade, resulting in a potent drug synergism that suppresses breast tumour growth. These findings highlight how metabolic redundancy can be context dependent, with the catalytic properties of different metabolic enzymes that act on the same substrate determining which pathways can support tumour growth in a particular nutrient environment. This, in turn, has practical consequences for therapies targeting cancer metabolism.

Epigenome-wide association study on the plasma metabolome suggests self-regulation of the glycine and serine pathway through DNA methylation.

BACKGROUND: The plasma metabolome reflects the physiological state of various biological processes and can serve as a proxy for disease risk. Plasma metabolite variation, influenced by genetic and epigenetic mechanisms, can also affect the cellular microenvironment and blood cell epigenetics. The interplay between the plasma metabolome and the blood cell epigenome remains elusive. In this study, we performed an epigenome-wide association study (EWAS) of 1183 plasma metabolites in 693 participants from the LifeLines-DEEP cohort and investigated the causal relationships in DNA methylation-metabolite associations using bidirectional Mendelian randomization and mediation analysis. RESULTS: After rigorously adjusting for potential confounders, including genetics, we identified five robust associations between two plasma metabolites (L-serine and glycine) and three CpG sites located in two independent genomic regions (cg14476101 and cg16246545 in PHGDH and cg02711608 in SLC1A5) at a false discovery rate of less than 0.05. Further analysis revealed a complex bidirectional relationship between plasma glycine/serine levels and DNA methylation. Moreover, we observed a strong mediating role of DNA methylation in the effect of glycine/serine on the expression of their metabolism/transport genes, with the proportion of the mediated effect ranging from 11.8 to 54.3%. This result was also replicated in an independent population-based cohort, the Rotterdam Study. To validate our findings, we conducted in vitro cell studies which confirmed the mediating role of DNA methylation in the regulation of PHGDH gene expression. CONCLUSIONS: Our findings reveal a potential feedback mechanism in which glycine and serine regulate gene expression through DNA methylation.

A handy way for forming N-doped TiO2/carbon from pectin and d,l-serine hydrazide hydrochloride.

N-doped TiO2/carbon composites (N-TiPC) have shown excellent photodegradation performances to the organic contaminants but are limited by the multistage preparation (i.e., preparation of porous carbon, preparation of N-doped TiO2, and loading of N-doped TiO2 on porous carbon). Here, we develop a handy way by combining the Pickering emulsion-gel template route and chelation reaction of polysaccharides. The N-TiPC is obtained by calcinating pectin/Dl-serine hydrazide hydrochloride (SHH)-Ti4+ chelate and is further described by modern characterization techniques. The results show that the N atom is successfully doped into the TiO2 lattice, and the bandgap value of N-TiPC is reduced to 2.3 eV. Moreover, the particle size of N-TiPC remains about 10 nm. The configurations of the composites are simulated using DFT calculation. The photocatalytic experiments show that N-TiPC has a high removal efficiency for methylene blue (MB) and oxytetracycline hydrochloride (OTC-HCL). The removal ratios of MB (20 mg/L, 50 mL) and OTC-HCL (30 mg/L, 50 mL) are 99.41 % and 78.29 %, respectively. The cyclic experiments show that the photocatalyst has good stability. Overall, this study provides a handy way to form N-TiPC with enhanced photodegradation performances. It can also be promoted to other macromolecules such as cellulose and its derivatives, sodium alginate, chitosan, lignin, etc.

An Immunometabolic Route for Activating cGAS/STING to Drive Anticancer Immunity.

The cGAS/STING pathway is a crucial immune activator in cancer biology, triggering innate immunosurveillance against tumors by sensing and reacting to endogenous mitochondrial DNA (mtDNA). In this issue of Cancer Research, research by Saha and colleagues highlights the significant impact of serine deprivation on this pathway, thereby unveiling its potential for anticancer therapy. Serine is essential for cellular metabolism and influences tumor growth and immune responses. Depriving cells of serine caused mitochondrial dysfunction and the release of mtDNA into the cytosol, activating the cGAS/STING pathway and inducing type I IFN responses. In mouse models, serine deprivation enhanced antitumor immunity, with increased tumoral immune infiltration, including CD4+/CD8+ T cells and type I IFN responses. Clinically, a genetic signature indicative of lower serine enrichment in colorectal cancer patients correlated with immune activation and improved survival. Furthermore, combining serine deprivation with PD1 blockade significantly reduced tumor volume and led to long-term immunity in mice, suggesting that serine depletion enhances the efficacy of immune checkpoint blockade. These findings propose serine deprivation as a promising strategy to boost antitumor immunity and improve cancer patient outcomes. See related article by Saha et al., p. 2645.

A lactate-dependent shift of glycolysis mediates synaptic and cognitive processes in male mice.

Astrocytes control brain activity via both metabolic processes and gliotransmission, but the physiological links between these functions are scantly known. Here we show that endogenous activation of astrocyte type-1 cannabinoid (CB1) receptors determines a shift of glycolysis towards the lactate-dependent production of D-serine, thereby gating synaptic and cognitive functions in male mice. Mutant mice lacking the CB1 receptor gene in astrocytes (GFAP-CB1-KO) are impaired in novel object recognition (NOR) memory. This phenotype is rescued by the gliotransmitter D-serine, by its precursor L-serine, and also by lactate and 3,5-DHBA, an agonist of the lactate receptor HCAR1. Such lactate-dependent effect is abolished when the astrocyte-specific phosphorylated-pathway (PP), which diverts glycolysis towards L-serine synthesis, is blocked. Consistently, lactate and 3,5-DHBA promoted the co-agonist binding site occupancy of CA1 post-synaptic NMDA receptors in hippocampal slices in a PP-dependent manner. Thus, a tight cross-talk between astrocytic energy metabolism and gliotransmission determines synaptic and cognitive processes.

Naturally Occurring Dehydrocostus Lactone Covalently Binds to KARRIKIN INSENSITIVE 2 by Dual Serine Modifications in Orobanche cumana and Arabidopsis.

Parasitic weeds, such as Orobanche and Striga, threaten crops globally. Contiguous efforts on the discovery and development of structurally novel seed germination stimulants targeting HYPOSENSITIVE TO LIGHT/KARRIKIN INSENSITIVE 2 (HTL/KAI2) have been made with the goal of weed control. Here, we demonstrate that a natural compound dehydrocostus lactone (DCL) exhibits effective "suicide germination" activity against Orobanche cumana and covalently binds to OcKAI2d2 on two catalytic serine sites with the second modification dependent on the first one. The same interactions and covalent modifications of DCL are also confirmed in AtKAI2. Further in-depth evolution analysis indicates that the proposed two catalytic sites are present throughout the streptophyte algae, hornworts, lycophytes, and seed plants. This discovery is particularly noteworthy as it signifies the first confirmation of a plant endogenous molecule directly binding to KAI2, which is valuable for unraveling the elusive identity of the KAI2 ligand and for targeting KAI2 paralogues for the development of novel germination stimulants.

Knockdown of ERN1 disturbs the expression of phosphoserine aminotransferase 1 and related genes in glioblastoma cells.

BACKGROUND: Endoplasmic reticulum stress and synthesis of serine are essential for tumor growth, but the mechanism of their interaction is not clarified yet. The overarching goal of this work was to investigate the impact of ERN1 (endoplasmic reticulum to nucleus signaling 1) inhibition on the expression of serine synthesis genes in U87MG glioblastoma cells concerning the suppression of cell proliferation. METHODS: Wild type U87MG glioblastoma cells and their clones with overexpression of transgenes dnERN1 (without cytoplasmic domain of ERN1) and dnrERN1 (with mutation in endoribonuclease of ERN1), and empty vector (as control) were used. The silencing of ERN1 and XBP1 was also used to inhibition of ERN1 and its function. Gene expression was measured by qPCR. RESULTS: We show that the expression of PSAT1 and several other related to serine synthesis genes is suppressed in cells with ERN1 inhibition by dissimilar mechanisms: PHGDH gene through ERN1 protein kinase, because its expression was resistant to inhibition of ERN1 endoribonuclease, but ATF4 gene via endoribonuclease of ERN1. However, in the control of PSAT1 and PSPH genes both enzymatic activities of ERN1 signaling protein are involved. At the same time, ERN1 knockdown strongly increased SHMT1 expression, which controls serine metabolism and enhances the proliferation and invasiveness of glioma cells. The level of microRNAs, which have binding sites in PSAT1, SHMT1, and PSPH mRNAs, was also changed in cells harboring dnERN1 transgene. Inhibition of ERN1 suppressed cell proliferation and enzymatic activity of PHGDH, a rate-limiting enzyme for serine synthesis. CONCLUSION: Changes in the expression of phosphoserine aminotransferase 1 and other genes related to serine synthesis are mediated by diverse ERN1-dependent mechanisms and contributed to suppressed proliferation and enhanced invasiveness of ERN1 knockdown glioblastoma cell.

ASCT2 is a major contributor to serine uptake in cancer cells.

The non-essential amino acid serine is a critical nutrient for cancer cells due to its diverse biosynthetic functions. While some tumors can synthesize serine de novo, others are auxotrophic and therefore reliant on serine uptake. Importantly, despite several transporters being known to be capable of transporting serine, the transporters that mediate serine uptake in cancer cells are not known. Here, we characterize the amino acid transporter ASCT2 (SLC1A5) as a major contributor to serine uptake in cancer cells. ASCT2 is well known as a glutamine transporter in cancer, and our work demonstrates that serine and glutamine compete for uptake through ASCT2. We further show that ASCT2-mediated serine uptake is essential for purine nucleotide biosynthesis and that estrogen receptor α (ERα) promotes serine uptake by directly activating SLC1A5 transcription. Collectively, our work defines an additional important role for ASCT2 as a serine transporter in cancer and evaluates ASCT2 as a potential therapeutic target.

Shifting Mycobacterial Serine Hydrolase Activity Visualized Using Multi-Layer In-Gel Activity Assays.

The ability of Mycobacterium tuberculosis to derive lipids from the host, store them intracellularly, and then break them down into energy requires a battery of serine hydrolases. Serine hydrolases are a large, diverse enzyme family with functional roles in dormant, active, and reactivating mycobacterial cultures. To rapidly measure substrate-dependent shifts in mycobacterial serine hydrolase activity, we combined a robust mycobacterial growth system of nitrogen limitation and variable carbon availability with nimble in-gel fluorogenic enzyme measurements. Using this methodology, we rapidly analyzed a range of ester substrates, identified multiple hydrolases concurrently, observed functional enzyme shifts, and measured global substrate preferences. Within every growth condition, mycobacterial hydrolases displayed the full, dynamic range of upregulated, downregulated, and constitutively active hydrolases independent of the ester substrate. Increasing the alkyl chain length of the ester substrate also allowed visualization of distinct hydrolase activity likely corresponding with lipases most responsible for lipid breakdown. The most robust expression of hydrolase activity was observed under the highest stress growth conditions, reflecting the induction of multiple metabolic pathways scavenging for energy to survive under this high stress. The unique hydrolases present under these high-stress conditions could represent novel drug targets for combination treatment with current front-line therapeutics. Combining diverse fluorogenic esters with in-gel activity measurements provides a rapid, customizable, and sensitive detection method for mycobacterial serine hydrolase activity.

Astrocytes enhance plasticity response during reversal learning.

Astrocytes play a key role in the regulation of synaptic strength and are thought to orchestrate synaptic plasticity and memory. Yet, how specifically astrocytes and their neuroactive transmitters control learning and memory is currently an open question. Recent experiments have uncovered an astrocyte-mediated feedback loop in CA1 pyramidal neurons which is started by the release of endocannabinoids by active neurons and closed by astrocytic regulation of the D-serine levels at the dendrites. D-serine is a co-agonist for the NMDA receptor regulating the strength and direction of synaptic plasticity. Activity-dependent D-serine release mediated by astrocytes is therefore a candidate for mediating between long-term synaptic depression (LTD) and potentiation (LTP) during learning. Here, we show that the mathematical description of this mechanism leads to a biophysical model of synaptic plasticity consistent with the phenomenological model known as the BCM model. The resulting mathematical framework can explain the learning deficit observed in mice upon disruption of the D-serine regulatory mechanism. It shows that D-serine enhances plasticity during reversal learning, ensuring fast responses to changes in the external environment. The model provides new testable predictions about the learning process, driving our understanding of the functional role of neuron-glia interaction in learning.