O-GlcNAcylation promotes topoisomerase IIα catalytic activity in breast cancer chemoresistance.

DNA topoisomerase IIα (TOP2A) plays a vital role in replication and cell division by catalytically altering DNA topology. It is a prominent target for anticancer drugs, but clinical efficacy is often compromised due to chemoresistance. In this study, we investigate the role of TOP2A O-GlcNAcylation in breast cancer cells and patient tumor tissues. Our results demonstrate that elevated TOP2A, especially its O-GlcNAcylation, promotes breast cancer malignant progression and resistance to adriamycin (Adm). O-GlcNAcylation at Ser1469 enhances TOP2A chromatin DNA binding and catalytic activity, leading to resistance to Adm in breast cancer cells and xenograft models. Mechanistically, O-GlcNAcylation-modulated interactions between TOP2A and cell cycle regulators influence downstream gene expression and contribute to breast cancer drug resistance. These results reveal a previously unrecognized mechanistic role for TOP2A O-GlcNAcylation in breast cancer chemotherapy resistance and provide support for targeting TOP2A O-GlcNAcylation in cancer therapy.

Dietary γ-mangostin triggers immunogenic cell death and activates cGAS signaling in acute myeloid leukemia.

Immunogenic cell death (ICD), one of cell-death types through release of damage-associated molecular patterns from dying tumor cells, activates tumor-specific immune response and elicits anti-tumor immunity by traditional radiotherapy and chemotherapy. However, whether natural products could induce ICD in leukemia is not elucidated. Here, we report dietary γ-mangostin eradicates murine primary leukemic cells and prolongs the survival of leukemic mice. As well, it restrains primary leukemic cells and CD34+ leukemic progenitor cells from leukemia patients. Strikingly, γ-mangostin attenuates leukemic cells by inducing ICD as characterized by expression of HSP90B1, ANXA1 and IL1B. Additionally, γ-mangostin accelerates cytoplasmic chromatin fragments generation, promoting DNA damage response, and enhances cGAS activation, leading to up-regulation of chemokines. Meanwhile, it induces HDAC4 degradation and acetylated histone H3 accumulation, which promotes chemokines transcription. Ultimately, CD8+ T cell is activated and recruited by γ-mangostin-induced chemokines in the microenvironment. Our study identifies γ-mangostin triggers ICD and activates cGAS signaling through DNA damage response and epigenetic modification. Therefore, dietary γ-mangostin would act as a potential agent to provoke anti-tumor immunity in the prevention and treatment of leukemia.

Cardiac magnetic resonance-derived myocardial scar is associated with echocardiographic response and clinical prognosis of left bundle branch area pacing for cardiac resynchronization therapy.

AIMS: Left bundle branch area pacing (LBBAP) is a novel approach for cardiac resynchronization therapy (CRT), but the impact of myocardial substrate on its effect is poorly understood. This study aims to assess the association of cardiac magnetic resonance (CMR)-derived scar burden and the response of CRT via LBBAP. METHODS AND RESULTS: Consecutive patients with CRT indications who underwent CMR examination and successful LBBAP-CRT were retrospectively analysed. Cardiac magnetic resonance late gadolinium enhancement was used for scar assessment. Echocardiographic reverse remodelling and composite outcomes (defined as all-cause death or heart failure hospitalization) were evaluated. The echocardiographic response was defined as a ≥15% reduction of left ventricular end-systolic volume. Among the 54 patients included, LBBAP-CRT resulted in a 74.1% response rate. The non-responders had higher global, septal, and lateral scar burden (all P < 0.001). Global, septal, and lateral scar percentage all predicted echocardiographic response [area under the curve (AUC): 0.857, 0.864, and 0.822; positive likelihood ratio (+LR): 9.859, 5.594, and 3.059; and negative likelihood ratio (-LR): 0.323, 0.233, and 0.175 respectively], which was superior to QRS morphology criteria (Strauss left bundle branch abnormality: AUC: 0.696, +LR 2.101, and -LR 0.389). After a median follow-up time of 20.3 (11.5-38.7) months, higher global, lateral and septal scar burdens were all predictive of the composite outcome (hazard ratios: 4.996, 7.019, and 4.741, respectively; P's < 0.05). CONCLUSION: Lower scar burden was associated with higher response rate of LBBAP-CRT. The pre-procedure CMR scar evaluation provides further useful information to identify potential responders and clinical outcomes.

The Cytotoxic Cardiac Glycoside (-)-Cryptanoside A from the Stems of Cryptolepis dubia and Its Molecular Targets.

A cardiac glycoside epoxide, (-)-cryptanoside A (1), was isolated from the stems of Cryptolepis dubia collected in Laos, for which the complete structure was confirmed by analysis of its spectroscopic and single-crystal X-ray diffraction data, using copper radiation at a low temperature. This cardiac glycoside epoxide exhibited potent cytotoxicity against several human cancer cell lines tested, including HT-29 colon, MDA-MB-231 breast, OVCAR3 and OVCAR5 ovarian cancer, and MDA-MB-435 melanoma cells, with the IC50 values found to be in the range 0.1-0.5 µM, which is comparable with that observed for digoxin. However, it exhibited less potent activity (IC50 1.1 µM) against FT194 benign/nonmalignant human fallopian tube secretory epithelial cells when compared with digoxin (IC50 0.16 µM), indicating its more selective activity toward human cancer versus benign/nonmalignant cells. (-)-Cryptanoside A (1) also inhibited Na+/K+-ATPase activity and increased the expression of Akt and the p65 subunit of NF-κB but did not show any effects on the expression of PI3K. A molecular docking profile showed that (-)-cryptanoside A (1) binds to Na+/K+-ATPase, and thus 1 may directly target Na+/K+-ATPase to mediate its cancer cell cytotoxicity.

The catalytic mechanism of direction-dependent interactions for 2,3-dihydroxybenzoate decarboxylase.

Benzoic acid decarboxylases offer an elegant alternative to CO2 fixation by reverse reaction-carboxylation, which is named the bio-Kolbe-Schmitt reaction, but they are unfavorable to carboxylation. Enhancing the carboxylation efficiency of reversible benzoic acid decarboxylases is restricted by the unexplained carboxylation mechanisms. The direction of reversible enzyme catalytic reactions depends on whether catalytic residues at the active center of the enzyme are protonated, which is subjected by the pH. Therefore, the forward and reverse reactions could be separated at different pH values. Reversible 2,3-dihydroxybenzoate acid decarboxylase undergoes decarboxylation at pH 5.0 and carboxylation at pH 8.6. However, it is unknown whether the interaction of enzymes with substrates and products in the forward and reverse reactions can be exploited to improve the catalytic activity of reversible enzymes in the unfavorable direction. Here, we identify a V-shaped tunnel of 2,3-dihydroxybenzoic acid decarboxylase from Aspergillus oryzae (2,3-DHBD_Ao) through which the substrate travels in the enzyme, and demonstrate that the side chain conformation of a tyrosine residue controls the entry and exit of substrate/product during reversible reactions. Together with the kinetic studies of the mutants, it is clarified that interactions between substrate/product traveling through the enzyme tunnel in 2,3-DHBD_Ao are direction-dependent. These results enrich the understanding of the interactions of substrates/products with macromolecular reversible enzymes in different reaction directions, thereby demonstrating a possible path for engineering decarboxylases with higher carboxylation efficiency. KEY POINTS: • The residue Trp23 of 2,3-DHBD_Ao served as a switch to control the entry and exit of catechol • A V-shaped tunnel of 2,3-DHBD_Ao for decarboxylation and carboxylation reactions was identified • The results provide a promising strategy for engineering decarboxylases with direction-dependent residues inside the substrate/product traveling tunnel of the enzyme.

A Targetless Method for Simultaneously Measuring Three-Degree-of-Freedom Angular Motion Errors with Digital Speckle Pattern Interferometry.

As a guide rail is the basic motion unit of precision equipment, the measurement of and compensation for its motion errors are important preconditions for precision machining and manufacturing. A targetless and simultaneous measurement method of three-degree-of-freedom (3-DOF) angular motion errors using digital speckle pattern interferometry (DSPI) is introduced in this paper. Based on the analysis of the sensitivity mechanism of DSPI to DOF errors and the formation mechanism of the phase fringes, the relationship between the angular motion errors and the distribution of the interferometric phases was established, and a new simultaneous measurement model of 3-DOF angular motion errors was further proposed. An optical setup based on a three-dimensional spatial-carrier DSPI with a right-angle symmetrical layout was used in the measurement system. Furthermore, repetitive tests, noise tests, and precision analysis were carried out to verify the performance of the system. The test results showed that the measurement resolution of the system was <1 µrad, which is capable of measuring the pitch angle, yaw angle, and roll angle at the submicron arc level simultaneously without target mirrors. The method has the advantages of no need to install cooperative targets and high measurement resolution, showing broad application prospects in many fields, including mechanical manufacturing, laser detection, aerospace, etc.

Variations in surface functional groups, carbon chemical state and graphitization degree during thermal deactivation of diesel soot particles.

The thermal deactivation of diesel soot particles exerts a significant influence on the control strategy for the regeneration of diesel particulate filters (DPFs). This work focused on the changes in the surface functional groups, carbon chemical state, and graphitization degree during thermal treatment in an inert gas environment at intermediate temperatures of 600°C, 800°C, and 1000°C and explore the chemical species that were desorbed from the diesel soot surface during thermal treatment using a thermogravimetric analyser coupled with a gas-chromatograph mass spectrometer (TGA-GC/MS). The surface functional groups and carbon chemical state were characterized using Fourier transform infrared spectroscopy (FT-IR) and X-ray photoelectron spectroscopy (XPS). The graphitization degree was evaluated by means of Raman spectroscopy (RS). The concentrations of aliphatic C-H, C-OH, C=O, and O-C=O groups are reduced for diesel soot and carbon black when increasing the thermal treatment temperature, while the sp2/sp3 hybridized ratio and graphitization degree enhance. These results provide comprehensive evidence of the decreased reactivity of soot samples. Among oxygenated functional groups, the percentage reduction during thermal treatment is the largest for the O-C=O groups owing to its worst thermodynamic stability. TGA-GC/MS results show that the aliphatic and aromatic chains and oxygenated species would be desorbed from the soot surface during 1000°C thermal treatment of diesel soot.

ppGalNAc-T4-catalyzed O-Glycosylation of TGF-ß type â ¡ receptor regulates breast cancer cells metastasis potential.

GalNAc-type O-glycosylation, initially catalyzed by polypeptide N-acetylgalactosaminyltransferases (ppGalNAc-Ts), is one of the most abundant and complex posttranslational modifications of proteins. Emerging evidence has proven that aberrant ppGalNAc-Ts are involved in malignant tumor transformation. However, the exact molecular functions of ppGalNAc-Ts are still unclear. Here, the role of one isoform, ppGalNAc-T4, in breast cancer cell lines was investigated. The expression of ppGalNAc-T4 was found to be negatively associated with migration of breast cancer cells. Loss-of-function studies revealed that ppGalNAc-T4 attenuated the migration and invasion of breast cancer cells by inhibiting the epithelial-mesenchymal transition (EMT) process. Correspondingly, transforming growth factor beta (TGF-ß) signaling, which is the upstream pathway of EMT, was impaired by ppGalNAc-T4 expression. ppGalNAc-T4 knockout decreased O-GalNAc modification of TGF-ß type Ⅰ and Ⅱ receptor (TßR Ⅰ and Ⅱ) and led to the elevation of TGF-ß receptor dimerization and activity. Importantly, a peptide from TßR Ⅱ was identified as a naked peptide substrate of ppGalNAc-T4 with a higher affinity than ppGalNAc-T2. Further, Ser31, corresponding to the extracellular domain of TßR Ⅱ, was identified as the O-GalNAcylation site upon in vitro glycosylation by ppGalNAc-T4. The O-GalNAc-deficient S31 A mutation enhanced TGF-ß signaling activity and EMT in breast cancer cells. Together, these results identified a novel mechanism of ppGalNAc-T4-catalyzed TGF-ß receptors O-GalNAcylation that suppresses breast cancer cell migration and invasion via the EMT process. Targeting ppGalNAc-T4 may be a potential therapeutic strategy for breast cancer treatment.

Targeting SOST using a small-molecule compound retards breast cancer bone metastasis.

BACKGROUND: Breast cancer metastasis to the bone can be exacerbated by osteoporosis, is associated with poor long-term survival, and has limited therapeutic options. Sclerostin (SOST) is an endogenous inhibitor of bone formation, and an attractive target for treatment of osteoporosis. However, it is unclear whether SOST can be used as a therapeutic target for bone metastases of breast cancer, and whether small molecule compounds that target SOST in breast cancer cells can inhibit breast cancer bone metastasis. METHODS: SOST expression in 442 breast cancer tissues was characterized by immunohistochemistry and statistically analyzed for the association with breast cancer bone metastases. Bone metastatic breast cancer SCP2 cells were induced for SOST silencing or overexpression and their bone metastatic behaviors were tested in vitro and in vivo. To identify potential therapeutics, we screened inhibitors of the interaction of SOST with STAT3 from a small chemical molecule library and tested the inhibitory effects of one inhibitor on breast cancer growth and bone metastasis in vitro and in vivo. RESULTS: We found that up-regulated SOST expression was associated with breast cancer bone metastases and worse survival of breast cancer patients. SOST silencing significantly reduced the bone metastatic capacity of SCP2 cells. SOST interacted with STAT3 to enhance the TGF-ß/KRAS signaling, increasing both tumor growth and bone metastasis. Treatment with one lead candidate, S6, significantly inhibited the growth of breast-cancer organoids and bone metastasis in mice. CONCLUSIONS: Our findings highlight a new class of potential therapeutics for treatment of bone metastasis in breast cancer.

Structural Insights into the Interactions of Digoxin and Na+/K+-ATPase and Other Targets for the Inhibition of Cancer Cell Proliferation.

Digoxin is a cardiac glycoside long used to treat congestive heart failure and found recently to show antitumor potential. The hydroxy groups connected at the C-12, C-14, and C-3'a positions; the C-17 unsaturated lactone unit; the conformation of the steroid core; and the C-3 saccharide moiety have been demonstrated as being important for digoxin's cytotoxicity and interactions with Na+/K+-ATPase. The docking profiles for digoxin and several derivatives and Na+/K+-ATPase were investigated; an additional small Asn130 side pocket was revealed, which could be useful in the design of novel digoxin-like antitumor agents. In addition, the docking scores for digoxin and its derivatives were found to correlate with their cytotoxicity, indicating a potential use of these values in the prediction of the cancer cell cytotoxicity of other cardiac glycosides. Moreover, in these docking studies, digoxin was found to bind to FIH-1 and NF-κB but not HDAC, IAP, and PI3K, suggesting that this cardiac glycoside directly targets FIH-1, Na+/K+-ATPase, and NF-κB to mediate its antitumor potential. Differentially, digoxigenin, the aglycon of digoxin, binds to HDAC and PI3K, but not FIH-1, IAP, Na+/K+-ATPase, and NF-κB, indicating that this compound may target tumor autophagy and metabolism to mediate its antitumor propensity.

Interaction of (+)-Strebloside and Its Derivatives with Na+/K+-ATPase and Other Targets.

Docking profiles for (+)-strebloside, a cytotoxic cardiac glycoside identified from Streblus asper, and some of its derivatives and Na+/K+-ATPase have been investigated. In addition, binding between (+)-strebloside and its aglycone, strophanthidin, and several of their other molecular targets, including FIH-1, HDAC, KEAP1 and MDM2 (negative regulators of Nrf2 and p53, respectively), NF-κB, and PI3K and Akt1, have been inspected and compared with those for digoxin and its aglycone, digoxigenin. The results showed that (+)-strebloside, digoxin, and their aglycones bind to KEAP1 and MDM2, while (+)-strebloside, strophanthidin, and digoxigenin dock to the active pocket of PI3K, and (+)-strebloside and digoxin interact with FIH-1. Thus, these cardiac glycosides could directly target HIF-1, Nrf2, and p53 protein-protein interactions, Na+/K+-ATPase, and PI3K to mediate their antitumor activity. Overall, (+)-strebloside seems more promising than digoxin for the development of potential anticancer agents.

Reversible inhibitor of CRM1 sensitizes glioblastoma cells to radiation by blocking the NF-κB signaling pathway.

BACKGROUND: Activation of nuclear factor-kappa B (NF-κΒ) through DNA damage is one of the causes of tumor cell resistance to radiotherapy. Chromosome region 1 (CRM1) regulates tumor cell proliferation, drug resistance, and radiation resistance by regulating the nuclear-cytoplasmic translocation of important tumor suppressor proteins or proto-oncoproteins. A large number of studies have reported that inhibition of CRM1 suppresses the activation of NF-κΒ. Thus, we hypothesize that the reversible CRM1 inhibitor S109 may induce radiosensitivity in glioblastoma (GBM) by regulating the NF-κΒ signaling pathway. METHODS: This study utilized the cell counting kit-8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU), and colony formation assay to evaluate the effect of S109 combined with radiotherapy on the proliferation and survival of GBM cells. The therapeutic efficacy of S109 combined with radiotherapy was evaluated in vivo to explore the therapeutic mechanism of S109-induced GBM radiosensitization. RESULTS: We found that S109 combined with radiotherapy significantly inhibited GBM cell proliferation and colony formation. By regulating the levels of multiple cell cycle- and apoptosis-related proteins, the combination therapy induced G1 cell cycle arrest in GBM cells. In vivo studies showed that S109 combined with radiotherapy significantly inhibited the growth of intracranial GBM and prolonged survival. Importantly, we found that S109 combined with radiotherapy promoted the nuclear accumulation of IκΒα, and inhibited phosphorylation of p65 and the transcriptional activation of NF-κΒ. CONCLUSION: Our findings provide a new therapeutic regimen for improving GBM radiosensitivity as well as a scientific basis for further clinical trials to evaluate this combination therapy.

Cytotoxic and non-cytotoxic cardiac glycosides isolated from the combined flowers, leaves, and twigs of Streblus asper.

A new non-cytotoxic [(+)-17ß-hydroxystrebloside (1)] and two known cytotoxic [(+)-3'-de-O-methylkamaloside (2) and (+)-strebloside (3)] cardiac glycosides were isolated and identified from the combined flowers, leaves, and twigs of Streblus asper collected in Vietnam, with the absolute configuration of 1 established from analysis of its ECD and NMR spectroscopic data and confirmed by computational ECD calculations. A new 14,21-epoxycardanolide (3a) was synthesized from 3 that was treated with base. A preliminary structure-activity relationship study indicated that the C-14 hydroxy group and the C-17 lactone unit and the established conformation are important for the mediation of the cytotoxicity of 3. Molecular docking profiles showed that the cytotoxic 3 and its non-cytotoxic analogue 1 bind differentially to Na+/K+-ATPase. Compound 3 docks deeply in the Na+/K+-ATPase pocket with a sole pose, and its C-10 formyl and C-5, C-14, and C-4' hydroxy groups may form hydrogen bonds with the side-chains of Glu111, Glu117, Thr797, and Arg880 of Na+/K+-ATPase, respectively. However, 1 fits the cation binding sites with at least three different poses, which all depotentiate the binding between 1 and Na+/K+-ATPase. Thus, 3 was found to inhibit Na+/K+-ATPase, but 1 did not. In addition, the cytotoxic and Na+/K+-ATPase inhibitory 3 did not affect glucose uptake in human lung cancer cells, against which it showed potent activity, indicating that this cardiac glycoside mediates its cytotoxicity by targeting Na+/K+-ATPase but not by interacting with glucose transporters.

Na+/K+-ATPase-Targeted Cytotoxicity of (+)-Digoxin and Several Semisynthetic Derivatives.

(+)-Digoxin (1) is a well-known cardiac glycoside long used to treat congestive heart failure and found more recently to show anticancer activity. Several known cardenolides (2-5) and two new analogues, (+)-8(9)-ß-anhydrodigoxigenin (6) and (+)-17-epi-20,22-dihydro-21α-hydroxydigoxin (7), were synthesized from 1 and evaluated for their cytotoxicity toward a small panel of human cancer cell lines. A preliminary structure-activity relationship investigation conducted indicated that the C-12 and C-14 hydroxy groups and the C-17 unsaturated lactone unit are important for 1 to mediate its cytotoxicity toward human cancer cells, but the C-3 glycosyl residue seems to be less critical for such an effect. Molecular docking profiles showed that the cytotoxic 1 and the noncytotoxic derivative 7 bind differentially to Na+/K+-ATPase. The HO-12ß, HO-14ß, and HO-3'aα hydroxy groups of (+)-digoxin (1) may form hydrogen bonds with the side-chains of Asp121 and Asn122, Thr797, and Arg880 of Na+/K+-ATPase, respectively, but the altered lactone unit of 7 results in a rotation of its steroid core, which depotentiates the binding between this compound and Na+/K+-ATPase. Thus, 1 was found to inhibit Na+/K+-ATPase, but 7 did not. In addition, the cytotoxic 1 did not affect glucose uptake in human cancer cells, indicating that this cardiac glycoside mediates its cytotoxicity by targeting Na+/K+-ATPase but not by interacting with glucose transporters.

Structural and enzymatic characterization of acetolactate decarboxylase from Bacillus subtilis.

Acetoin is an important physiological metabolite excreted by microbes. Its functions include avoiding acidification, participating in regulation of the NAD+/NADH ratio, and storing carbon. Acetolactate decarboxylase is a well-characterized anabolic enzyme involved with 3-hydroxy butanone (acetoin). It catalyzes conversion of the (R)- and (S)-enantiomers of acetolactate to generate the single product, (R)-acetoin. In addition to the X-ray crystal structure of acetolactate decarboxylase from Bacillus brevis, although the enzyme is widely present in microorganisms, very few atomic structures of acetolactate decarboxylase are reported. In this paper, we solved and reported a 1.5 Å resolution crystal structure of acetolactate decarboxylase from Bacillus subtilis. Dimeric assembly is observed in the solved structure, which is consistent with the elution profile conducted by molecular filtration. A zinc ion is coordinated by highly conserved histidines (191, 193, and 204) and conserved glutamates (62 and 251). We performed kinetic studies on acetolactate decarboxylase from Bacillus subtilis using circular dichroism, allowing the conversion of acetolactate to chiral acetoin for real-time tracking, yielding a Km value of 21 mM and a kcat value of 2.2 s-1. Using the two enantiomers of acetolactate as substrates, we further investigated the substrate preference of acetolactate decarboxylase from Bacillus subtilis by means of molecular docking and dynamic simulation in silico. The binding free energy of (S)-acetolactate was found to be ~ 30 kcal/mol greater than that of (R)-acetolactate, indicating a more stable binding for (S)-acetolactate.

Spatial phase-shift dual-beam speckle interferometry.

The spatial phase-shift technique has been successfully applied to an out-of-plane speckle interferometry system. Its application to a pure in-plane sensitive system has not been reported yet. This paper presents a novel optical configuration that enables the application of the spatial phase-shift technique to pure in-plane sensitive dual-beam speckle interferometry. The new spatial phase-shift dual-beam speckle interferometry (SPS-DBSP) uses a dual-beam in-plane electronic speckle pattern interferometry configuration with individual aperture shears, avoiding the interference in the object plane by the use of a low-coherence source, and different optical paths. The measured object is illuminated by two incoherent beams that are generated by a delay line, which is larger than the coherence length of the laser. The two beams reflected from the object surface interfere with each other at the CCD plane because of different optical paths. A spatial phase shift is introduced by the angle between the two apertures when they are mapped to the same optical axis. The phase of the in-plane deformation can directly be extracted from the speckle patterns by the Fourier transform method. The capability of SPS-DBSI is demonstrated by theoretical discussion as well as experiments.

Identification and Characterization of the First Cathelicidin from Sea Snakes with Potent Antimicrobial and Anti-inflammatory Activity and Special Mechanism.

Cathelicidins are a family of gene-encoded peptide effectors of innate immunity found exclusively in vertebrates. They play pivotal roles in host immune defense against microbial invasions. Dozens of cathelicidins have been identified from several vertebrate species. However, no cathelicidin from marine reptiles has been characterized previously. Here we report the identification and characterization of a novel cathelicidin (Hc-CATH) from the sea snake Hydrophis cyanocinctus. Hc-CATH is composed of 30 amino acids, and the sequence is KFFKRLLKSVRRAVKKFRKKPRLIGLSTLL. Circular dichroism spectroscopy and structure modeling analysis indicated that Hc-CATH mainly assumes an amphipathic α-helical conformation in bacterial membrane-mimetic solutions. It possesses potent broad-spectrum and rapid antimicrobial activity. Meanwhile, it is highly stable and shows low cytotoxicity toward mammalian cells. The microbial killing activity of Hc-CATH is executed through the disruption of cell membrane and lysis of bacterial cells. In addition, Hc-CATH exhibited potent anti-inflammatory activity by inhibiting the LPS-induced production of nitric oxide (NO) and pro-inflammatory cytokines such as TNF-α, IL-1ß, and IL-6. Hc-CATH directly binds with LPS to neutralize its toxicity, and it also binds to Toll-like receptor 4 (TLR4/MD2 complex), which therefore inhibits the binding of LPS to TLR4/MD2 complex and the subsequent activation of LPS-induced inflammatory response pathways. Taken together, our study demonstrates that Hc-CATH, the first cathelicidin from sea snake discovered to have both antimicrobial and anti-inflammatory activity, is a potent candidate for the development of peptide antibiotics.

Cysteinome: The first comprehensive database for proteins with targetable cysteine and their covalent inhibitors.

The covalent modification of intrinsically nucleophilic cysteine in proteins is crucial for diverse biochemical events. Bioinformatics approaches may prove useful in the design and discovery of covalent molecules targeting the cysteine in proteins to tune their functions and activities. Herein, we describe the Cysteinome, the first online database that provides a rich resource for the display, search and analysis of structure, function and related annotation for proteins with targetable cysteine as well as their covalent modulators. To this end, Cysteinome compiles 462 proteins with targetable cysteine from 122 different species along with 1217 covalent modulators curated from existing literatures. Proteins are annotated with a detailed description of protein families, biological process and related diseases. In addition, covalent modulators are carefully annotated with chemical name, chemical structure, binding affinity, physicochemical properties, molecule type and related diseases etc. The Cysteinome database may serve as a useful platform for the identification of crucial proteins with targetable cysteine in certain cellular context. Furthermore, it may help biologists and chemists for the design and discovery of covalent chemical probes or inhibitors homing at functional cysteine of critical protein targets implicated in various physiological or disease process. The Cysteinome database is freely available to public at http://www.cysteinome.org/.

TmcN is involved in ATP regulation of tautomycetin biosynthesis in Streptomyces griseochromogenes.

The regulatory mechanism of tautomycetin (TMC) biosynthesis remains largely unknown, although it has been of great interest to the pharmaceutical industry. Our previous study showed that intracellular adenosine triphosphate (inATP) level is negatively correlated with secondary metabolite biosynthesis in various Streptomyces spp. In this study, by exogenous treatment of ATP, we also found a negative correlation between TMC biosynthesis and inATP level in Streptomyces griseochromogenes (S. griseochromogenes). However, the underlying mechanism remains unclear. TmcN, a pathway-specific transcriptional regulator of TMC biosynthetic genes, was previously revealed as a large ATP-binding LuxR (LAL) family protein. The predicted amino acid sequence of TmcN shows highly conserved Walker A and B binding motifs, which suggest an ATPase function of TmcN. We therefore hypothesized that the ATPase domain of TmcN may play a role in sensing endogenous pool of ATP, and is thus involved in the ATP regulation of TMC biosynthesis. To test the hypothesis, we first explored the key residue that affects the ATPase activity of TmcN by amino acid sequence alignment and structural simulation. After that, we disrupted tmcN gene in S. griseochromogenes, and the tmcN or site-direct-mutated tmcN were re-introduced to get the complementary and ATPase domain disrupted strains. The transcription level of tmcN, TMC yield, and inATP, as well as the effect of ATP on TMC production of different mutants were evaluated. Deletion of tmcN or site-direct mutation of ATPase domain of TmcN in S. griseochromogenes significantly reduced the TMC production, and it was not affected by exogenous ATP treatment. In addition, a relatively high level of inATP was detected in tmcN deletion and site-direct mutation strains. Our results here suggested that TmcN, especially its ATPase domain, is involved in consuming of endogenous ATP pool and thus plays pivotal role in connecting the primary and secondary metabolite in S. griseochromogenes.

Spatiotemporal three-dimensional phase unwrapping in digital speckle pattern interferometry.

We propose a hybrid spatiotemporal three-dimensional phase unwrapping algorithm for use in digital speckle pattern interferometry (DSPI). The feature of the proposed algorithm is the integration of one-dimensional temporal and two-dimensional spatial phase unwrapping algorithms. By demodulating the phase on a single reference point or multiple reference points using temporal phase unwrapping and on each separated phase map region using spatial phase unwrapping, the DSPI with the spatiotemporal three-dimensional phase unwrapping algorithm can realize the measurement of dynamic absolute displacements and the determination of abrupt phase changes which are usually caused by object discontinuities. We demonstrate that the presented algorithm can overcome the drawbacks of the traditional spatial and temporal phase unwrapping algorithms.