The SMC5/6 complex prevents genotoxicity upon APOBEC3A-mediated replication stress.

Mutational patterns caused by APOBEC3 cytidine deaminase activity are evident throughout human cancer genomes. In particular, the APOBEC3A family member is a potent genotoxin that causes substantial DNA damage in experimental systems and human tumors. However, the mechanisms that ensure genome stability in cells with active APOBEC3A are unknown. Through an unbiased genome-wide screen, we define the Structural Maintenance of Chromosomes 5/6 (SMC5/6) complex as essential for cell viability when APOBEC3A is active. We observe an absence of APOBEC3A mutagenesis in human tumors with SMC5/6 dysfunction, consistent with synthetic lethality. Cancer cells depleted of SMC5/6 incur substantial genome damage from APOBEC3A activity during DNA replication. Further, APOBEC3A activity results in replication tract lengthening which is dependent on PrimPol, consistent with re-initiation of DNA synthesis downstream of APOBEC3A-induced lesions. Loss of SMC5/6 abrogates elongated replication tracts and increases DNA breaks upon APOBEC3A activity. Our findings indicate that replication fork lengthening reflects a DNA damage response to APOBEC3A activity that promotes genome stability in an SMC5/6-dependent manner. Therefore, SMC5/6 presents a potential therapeutic vulnerability in tumors with active APOBEC3A.

The FLIP-FIGNL1 complex regulates the dissociation of RAD51/DMC1 in homologous recombination and replication fork restart.

Recruitment of RAD51 and/or DMC1 recombinases to single-strand DNA is indispensable for homology search and strand invasion in homologous recombination (HR) and for protection of nascent DNA strands at stalled replication forks. Thereafter RAD51/DMC1 dissociate, actively or passively, from these joint molecules upon DNA repair or releasing from replication stress. However, the mechanism that regulates RAD51/DMC1 dissociation and its physiological importance remain elusive. Here, we show that a FLIP-FIGNL1 complex regulates RAD51 and DMC1 dissociation to promote meiotic recombination and replication fork restart in mammals. Mice lacking FLIP are embryonic lethal, while germline-specific deletion of FLIP leads to infertility in both males and females. FLIP-null meiocytes are arrested at a zygotene-like stage with massive RAD51 and DMC1 foci, which frequently co-localize with SHOC1 and TEX11. Furthermore, FLIP interacts with FIGNL1. Depletion of FLIP or FIGNL1 in cell lines destabilizes each other and impairs RAD51 dissociation. Thus, the active dissociation of RAD51/DMC1 by the FLIP-FIGNL1 complex is a crucial step required for HR and replication fork restart, and represents a conserved mechanism in somatic cells and germ cells.

Internal Standard Assisted Surface-Enhanced Raman Scattering Nanoprobe with 4-NTP as Recognition Unit for Ratiometric Imaging Hydrogen Sulfide in Living Cells.

Hydrogen sulfide (H2S), as the third endogenous gasotransmitter, is closely associated with various physiological and pathological processes, whereas many aspects of its functions remain unclear. Effective tools for the accurate detection of H2S in living organisms are urgently needed. We herein reported an internal standard assisted surface-enhanced Raman scattering (SERS) nanoprobe for ratiometric detection of H2S in vitro and in living cells based on the reduction of nitros with H2S. This nanoprobe consists of an internal standard (4-mercaptobenzonitrile, MPBN) embedded core-molecule-shell Au nanoflower (Au@MPBN@Au) as the high plasmonic active SERS substrate and the 4-nitrothiophenol (4-NTP) molecule immobilized on the surface as the H2S recognition unit. With the addition of H2S, the nitros peak (1329 cm-1) decreased. Meanwhile, three obvious new peaks appeared at 1139, 1387, and 1433 cm-1, which were related to the vibration of the dimerized product 4,4'-dimercaptoazobisbenzene (DMAB) of 4-aminothiophenol (4-ATP). However, the peak intensity at 2223 cm-1 derived from MPBN was not influenced by the outer environment. Thus, the H2S level was able to be determined based on the ratio of two peak intensities (I1139/I2223) with a detection limit as low as 0.24 µM. Notably, we have proved that SERS nanoprobe Au@MPBN@Au@4-NTP could ratiometrically image both the endogenous and exogenous H2S in living cells. We anticipate that Au@MPBN@Au@4-NTP could be applied for the study of H2S-related physiological function in the future.

A near-infrared fluorescent probe for ratiometric sensing of SO2 in cells and zebrafish.

Developing suitable molecular tools for monitoring SO2 and its derivatives in living organisms is attractive due to their importance for human health. Herein, the first near-infrared fluorescent probe Mito-HN with aggregation-induced emission enhancement (AIEE) characteristics for ratiometric sensing of SO2 derivatives in vitro, in cells, and in zebrafish was designed and synthesized. By connecting the electron-donating naphthopyran and electron-withdrawing hemicyanine via the alkenyl group, this probe displays a near-infrared fluorescence emission and notable solid-state luminescence with AIEE features. Based on the specific 1,4-addition reaction of HSO3-/SO32- with the C[double bond, length as m-dash]C double bond of Mito-HN, this probe can specifically sense HSO3- in real time with an excellent near-infrared ratiometric fluorescence effect, a low detection limit of 0.17 µM, and a fast response time of less than 30 s. Cell imaging and zebrafish imaging results demonstrate that Mito-HN can ratiometrically not only sense endogenous SO2 derivatives in mitochondria but also monitor SO2 in living organisms. We anticipate that Mito-HN could be applied for the diagnosis of SO2-related diseases in the future.

LeuRS can leucylate type I and type II tRNALeus in Streptomyces coelicolor.

Transfer RNAs (tRNAs) are divided into two types, type I with a short variable loop and type II with a long variable loop. Aminoacylation of type I or type II tRNALeu is catalyzed by their cognate leucyl-tRNA synthetases (LeuRSs). However, in Streptomyces coelicolor, there are two types of tRNALeu and only one LeuRS (ScoLeuRS). We found that the enzyme could leucylate both types of ScotRNALeu, and had a higher catalytic efficiency for type II ScotRNALeu(UAA) than for type I ScotRNALeu(CAA). The results from tRNA and enzyme mutagenesis showed that ScoLeuRS did not interact with the canonical discriminator A73. The number of nucleotides, rather than the type of base of the variable loop in the two types of ScotRNALeus, was determined as important for aminoacylation. In vitro and in vivo assays showed that the tertiary structure formed by the D-loop and TψC-loop is more important for ScotRNALeu(UAA). We showed that the leucine-specific domain (LSD) of ScoLeuRS could help LeuRS, which originally only leucylates type II tRNALeu, to aminoacylate type I ScotRNALeu(CAA) and identified the crucial amino acid residues at the C-terminus of the LSD to recognize type I ScotRNALeu(CAA). Overall, our findings identified a rare recognition mechanism of LeuRS to tRNALeu.

The HIRA histone chaperone complex subunit UBN1 harbors H3/H4- and DNA-binding activity.

The HIRA histone chaperone complex is composed of the proteins HIRA, UBN1, and CABIN1 and cooperates with the histone chaperone ASF1a to specifically bind and deposit H3.3/H4 into chromatin. We recently reported that the UBN1 Hpc2-related domain (HRD) specifically binds to H3.3/H4 over H3.1/H4. However, the mechanism for HIRA complex deposition of H3.3/H4 into nucleosomes remains unclear. Here, we characterize a central region of UBN1 (UBN1 middle domain) that is evolutionarily conserved and predicted to have helical secondary structure. We report that the UBN1 middle domain has dimer formation activity and binds to H3/H4 in a manner that does not discriminate between H3.1 and H3.3. We additionally identify a nearby DNA-binding domain in UBN1, located between the UBN1 HRD and middle domain, which binds DNA through electrostatic contacts involving several conserved lysine residues. Together, these observations suggest a mechanism for HIRA-mediated H3.3/H4 deposition whereby UBN1 associates with DNA and dimerizes to mediate formation of an (H3.3/H4)2 heterotetramer prior to chromatin deposition.

Pharmacokinetic Characterization of (Poly)phenolic Metabolites in Human Plasma and Urine after Acute and Short-Term Daily Consumption of Mango Pulp.

Pharmacokinetic (PK) evaluation of polyphenolic metabolites over 24 h was conducted in human subjects (n = 13, BMI = 22.7 ± 0.4 kg/m2) after acute mango pulp (MP), vitamin C (VC) or MP + VC test beverage intake and after 14 days of MP beverage intake. Plasma and urine samples were collected at different time intervals and analyzed using targeted and non-targeted mass spectrometry. The maximum concentrations (Cmax) of gallotannin metabolites were significantly increased (p < 0.05) after acute MP beverage intake compared to VC beverage alone. MP + VC beverage non-significantly enhanced the Cmax of gallic acid metabolites compared to MP beverage alone. Pyrogallol (microbial-derived metabolite) derivatives increased (3.6%) after the 14 days of MP beverage intake compared to 24 h acute MP beverage intake (p < 0.05). These results indicate extensive absorption and breakdown of gallotannins to galloyl and other (poly)phenolic metabolites after MP consumption, suggesting modulation and/or acclimation of gut microbiota to daily MP intake.

Structure of the α-tubulin acetyltransferase, αTAT1, and implications for tubulin-specific acetylation.

Protein acetylation is an important posttranslational modification with the recent identification of new substrates and enzymes, new links to disease, and modulators of protein acetylation for therapy. α-Tubulin acetyltransferase (αTAT1) is the major α-tubulin lysine-40 (K40) acetyltransferase in mammals, nematodes, and protozoa, and its activity plays a conserved role in several microtubule-based processes. Here, we present the X-ray crystal structure of the human αTAT1/acetyl-CoA complex. Together with structure-based mutagenesis, enzymatic analysis, and functional studies in cells, we elucidate the catalytic mechanism and mode of tubulin-specific acetylation. We find that αTAT1 has an overall fold similar to the Gcn5 histone acetyltransferase but contains a relatively wide substrate binding groove and unique structural elements that play important roles in α-tubulin-specific acetylation. Conserved aspartic acid and cysteine residues play important catalytic roles through a ternary complex mechanism. αTAT1 mutations have analogous effects on tubulin acetylation in vitro and in cells, demonstrating that it is the central determining factor of α-tubulin K40 acetylation levels in vivo. Together, these studies provide general insights into distinguishing features between histone and tubulin acetyltransferases, and they have specific implications for understanding the molecular basis of tubulin acetylation and for developing small molecule modulators of microtubule acetylation for therapy.

The growth and medicinal quality of Epimedium wushanense are improved by an isolate of dark septate fungus.

CONTEXT: Seven dark-septate endophytic (DSE) fungi have been isolated from the roots of Epimedium wushanense T. S. Ying (Berberidaceae), an important medicinal plant with various pharmacological activities. OBJECTIVE: The current study explores the effects of seven DSE fungi on the growth and accumulation of bioactive compounds in E. wushanense. MATERIALS AND METHODS: Each 1-month-old E. wushanense seedling was inoculated with one of the seven DSE fungi and was grown under greenhouse conditions for 90 d. The molecular identification of the fungi was based on the ITS1-5.8S-ITS2 nuclear ribosomal gene cluster. RESULTS: The results showed that the influence of DSE fungi inoculation varied between strains. Inoculation with DSE8 not only significantly enhanced plant height, root length, leaf area, leaf number, and shoot and root biomass but also improved the total flavonoid and icariin content, with an increase ranging from 20.24% to 237.97%. Three of the seven DSE fungi caused the inoculated plants to die, and the remaining three DSE strains showed neutral or negative effects on plant growth and the accumulation of bioactive compounds. According to the ITS sequence, DSE8 is congeneric to the genus Leptodontidium. DISCUSSION AND CONCLUSION: The findings indicate that application of DSE8 may be valuable to facilitate the cultivation of E. wushanense with a higher biomass and improved medicinal quality.

Comparative analysis of EPA and DHA in fish oil nutritional capsules by GC-MS.

BACKGROUND: Fish oil is a popular nutritional product consumed in Hong Kong. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are the two main bioactive components responsible for the health benefits of fish oil. Market survey in Hong Kong demonstrated that various fish oil capsules with different origins and prices are sold simultaneously. However, these capsules are labelled with same ingredient levels, namely EPA 180 mg/g and DHA 120 mg/g. This situation makes the consumers very confused. To evaluate the quality of various fish oil capsules, a comparative analysis of the contents of EPA and DHA in fish oil is crucial. METHODS: A gas chromatography-mass spectrometry (GC-MS) method was developed for identification and determination of EPA and DHA in fish oil capsules. A comprehensive validation of the developed method was conducted. Ten batches of fish oil capsules samples purchased from drugstores of Hong Kong were analyzed by using the developed method. RESULTS: The present method presented good sensitivity, precision and accuracy. The limits of detection (LOD) for EPA and DHA were 0.08 ng and 0.21 ng, respectively. The relative standard deviation (RSD) values of EPA and DHA for repeatability tests were both less than 1.05%; and the recovery for accuracy test of EPA and DHA were 100.50% and 103.83%, respectively. In ten fish oil samples, the contents of EPA ranged from 39.52 mg/g to 509.16 mg/g, and the contents of DHA ranged from 35.14 mg/g to 645.70 mg/g. CONCLUSION: The present method is suitable for the quantitative analysis of EPA and DHA in fish oil capsules. There is a significant variation in the contents of the quantified components in fish oil samples, and there is not a linear relationship between price and contents of EPA and DHA. Strict supervision of the labelling of the fish oil capsules is urgently needed.

A systematic review of the botanical, phytochemical and pharmacological profile of Dracaena cochinchinensis, a plant source of the ethnomedicine "dragon's blood".

"Dragon's blood" is the name given to a deep red resin obtained from a variety of plant sources. The resin extracted from stems of Dracaena cochinchinensis is one such source of "dragon's blood". It has a reputation for facilitating blood circulation and dispersing blood stasis. In traditional Chinese medicine, this resinous medicine is commonly prescribed to invigorate blood circulation for the treatment of traumatic injuries, blood stasis and pain. Modern pharmacological studies have found that this resinous medicine has anti-bacterial, anti-spasmodic, anti-inflammatory, analgesic, anti-diabetic, and anti-tumor activities, while it is also known to enhance immune function, promote skin repair, stop bleeding and enhance blood circulation. Various compounds have been isolated from the plant, including loureirin A, loureirin B, loureirin C, cochinchinenin, socotrin-4'-ol, 4',7-dihydroxyflavan, 4-methylcholest-7-ene-3-ol, ethylparaben, resveratrol, and hydroxyphenol. The present review summarizes current knowledge concerning the botany, phytochemistry, pharmacological effects, toxicology studies and clinical applications of this resinous medicine as derived from D. cochinchinenesis.

Srs2 binding to PCNA and its sumoylation contribute to RPA antagonism during the DNA damage response.

Activation of the DNA damage checkpoint upon genotoxin treatment induces a multitude of cellular changes, such as cell cycle arrest, to cope with genome stress. After prolonged genotoxin treatment, the checkpoint can be downregulated to allow cell cycle and growth resumption. In yeast, downregulation of the DNA damage checkpoint requires the Srs2 DNA helicase, which removes the ssDNA binding complex RPA and the associated Mec1 checkpoint kinase from DNA, thus dampening Mec1 activation. However, it is unclear whether the 'anti-checkpoint' role of Srs2 is temporally and spatially regulated to both allow timely checkpoint termination and to prevent superfluous RPA removal. Here we address this question by examining regulatory elements of Srs2, including its phosphorylation, sumoylation, and protein-interaction sites. Our genetic analyses and checkpoint level assessment suggest that the RPA countering role of Srs2 is promoted by Srs2 binding to PCNA, which is known to recruit Srs2 to subsets of ssDNA regions. RPA antagonism is further fostered by Srs2 sumoylation, which we found depends on the Srs2-PCNA interaction. Srs2 sumoylation is additionally reliant on Mec1 and peaks after Mec1 activity reaches maximal levels. Collectively, our data provide evidence for a two-step model wherein checkpoint downregulation is facilitated by PCNA-mediated Srs2 recruitment to ssDNA-RPA filaments and the subsequent Srs2 sumoylation stimulated upon Mec1 hyperactivation. We propose that this mechanism allows Mec1 hyperactivation to trigger checkpoint recovery.

Chemical components with antibacterial properties found in sanchen powder from traditional Tibetan medicine.

ETHNOPHARMACOLOGICAL RELEVANCE: Sanchen powder is a traditional Tibetan medicine comprising Bambusae Concretio Silicea, Carthami Flos, and Bovis Calculus Artifactus. Bambusae Concretio Silicea is the dried mass of secreted fluid in the stalks of Gramineae plants such as Bambusa textilis McClure or Schizostachyum chinense Rendle. Carthami Flos is the dried flower of Carthamus tinctorius L. in the Compositae plant. Bovis Calculus Artifactus is made from ox bile powder, cholic acid, hyodeoxycholic acid, taurine, bilirubin, cholesterol, and trace elements. Research has evidenced the antibacterial efficacy of Sanchen powder, albeit its active constituents for this effect are yet to be established. AIM OF THE STUDY: To investigate effective compounds, potential targets, and molecular mechanism of Sanchen powder for its antibacterial properties by using network pharmacology combined with in vitro validation, with the aims of observing the action of effective compounds in Sanchen powder and exploring new therapeutic strategies for antibacterial. MATERIALS AND METHODS: In this study, UPLC-Q-TOF-MS was utilized to identify the chemical composition in Sanchen powder and its blood-borne chemical ingredients post-oral intake. A network pharmacology analysis was used to establish the chemical compound in the blood following oral administration-target-disease network. The study aimed to identify antibacterial active ingredients, which were then subjected to molecular docking and pharmacodynamic experiments to verify their efficacy. RESULTS: The findings demonstrate that following oral administration, the blood contains seven key components of Sanchen powder, including bilirubin, glycochenodeoxycholic acid, glycocholic acid, taurocholic acid, phenylalanine, safflomin A, and tryptophan. Additionally, the network pharmacology and molecular docking study results indicate the potential antibacterial effects of bilirubin, glycocholic acid, and glycochenodeoxycholic acid. In vitro antibacterial experiments revealed that bilirubin, glycocholic acid, and glycochenodeoxycholic acid could restrict the growth of the Staphylococcus aureus cell membrane at a certain concentration. Moreover, they exhibited antibacterial effects on Staphylococcus aureus, Pseudomonas aeruginosa, Staphylococcus epidermidis, and Escherichia coli. CONCLUSIONS: Bilirubin, glycocholic acid, and glycochenodeoxycholic acid could be effective therapeutic ingredients for the antibacterial effects of Sanchen powder. These results offer a foundation for further clinical application and research on the antibacterial effect of Sanchen powder, a Traditional Tibetan Medicine.

Effects of micro-nano plastics on the environmental biogeochemical cycle of nitrogen: A comprehensive review.

Micro-nano plastics (MNPs; size <5 mm), ubiquitous and emerging pollutants, accumulated in the natural environment through various sources, and are likely to interact with nutrients, thereby influencing their biogeochemical cycle. Increasing scientific evidences reveal that MNPs can affect nitrogen (N) cycle processes by affecting biotopes and organisms in the environmental matrix and MNPs biofilms, thus plays a crucial role in nitrous oxide (N2O) and ammonia (NH3) emission. Yet, the mechanism and key processes behind this have not been systematically reviewed in natural environments. In this review, we systematically summarize the effects of MNPs on N transformation in terrestrial, aquatic, and atmospheric ecosystems. The effects of MNPs properties on N content, composition, and function of the microbial community, enzyme activity, gene abundance and plant N uptake in different environmental conditions has been briefly discussed. The review highlights the significant potential of MNPs to alter the properties of the environmental matrix, microbes and plant or animal physiology, resulting in changes in N uptake and metabolic efficiency in plants, thereby inhibiting organic nitrogen (ON) formation and reducing N bioavailability, or altering NH3 emissions from animal sources. The faster the decomposition of plastics, the more intense the perturbation of MNPs to organisms in the natural ecosystem. Findings of this provide a more comprehensive analysis and research directions to the environmentalists, policy makers, water resources planners & managers, biologists, and biotechnologists to do integrate approaches to reach the practical engineering solutions which will further diminish the long-term ecological and climatic risks.

Expanding Complex Morpholines Using Systematic Chemical Diversity.

The morpholine heterocycle is a structural unit found in many bioactive compounds and FDA-approved drugs, but the generation of more complex C-functionalized morpholine derivatives remains considerably underexplored. Using systematic chemical diversity (SCD), a concept that guides the expansion of saturated drug-like scaffolds through regiochemical and stereochemical variation, we describe the synthesis of a collection of methyl-substituted morpholine acetic acid esters starting from enantiomerically pure amino acids and amino alcohols. In total, 24 diverse substituted morpholines were produced that vary systematically in regiochemistry and stereochemistry (relative and absolute). These diverse C-substituted morpholines can be directly applied in fragment screening or incorporated as building blocks in medicinal chemistry and library synthesis.

Overexpression of soybean GmDHN9 gene enhances drought resistance of transgenic Arabidopsis.

Soybean is one of the important oil crops and a major source of protein and lipids. Drought can cause severe soybean yields. Dehydrin protein (DHN) is a subfamily of LEA proteins that play an important role in plant responses to abiotic stresses. In this study, the soybean GmDHN9 gene was cloned and induced under a variety of abiotic stresses. Results showed that the GmDHN9 gene response was more pronounced under drought induction. Subcellular localization results indicated that the protein was localized in the cytoplasm. The role of transgenic Arabidopsis plants in drought stress response was further studied. Under drought stress, the germination rate, root length, chlorophyll, proline, relative water content, and antioxidant enzyme content of transgenic Arabidopsis thaliana transgenic genes were higher than those of wild-type plants, and transgenic plants contained less O2-, H2O2 and MDA contents. In short, the GmDHN9 gene can regulate the homeostasis of ROS and enhance the drought resistance of plants.

Exploiting the Carboxylate-Binding Pocket of ß-Lactamase Enzymes Using a Focused DNA-Encoded Chemical Library.

ß-Lactamase enzymes hydrolyze and thereby provide bacterial resistance to the important ß-lactam class of antibiotics. The OXA-48 and NDM-1 ß-lactamases cause resistance to the last-resort ß-lactams, carbapenems, leading to a serious public health threat. Here, we utilized DNA-encoded chemical library (DECL) technology to discover novel ß-lactamase inhibitors. We exploited the ß-lactamase enzyme-substrate binding interactions and created a DECL targeting the carboxylate-binding pocket present in all ß-lactamases. A library of 106 compounds, each containing a carboxylic acid or a tetrazole as an enzyme recognition element, was designed, constructed, and used to identify OXA-48 and NDM-1 inhibitors with micromolar to nanomolar potency. Further optimization led to NDM-1 inhibitors with increased potencies and biological activities. This work demonstrates that the carboxylate-binding pocket-targeting DECL, designed based on substrate binding information, aids in inhibitor identification and led to the discovery of novel non-ß-lactam pharmacophores for the development of ß-lactamase inhibitors for enzymes of different structural and mechanistic classes.

The SMC5/6 complex prevents genotoxicity upon APOBEC3A-mediated replication stress.

Mutational patterns caused by APOBEC3 cytidine deaminase activity are evident throughout human cancer genomes. In particular, the APOBEC3A family member is a potent genotoxin that causes substantial DNA damage in experimental systems and human tumors. However, the mechanisms that ensure genome stability in cells with active APOBEC3A are unknown. Through an unbiased genome-wide screen, we define the Structural Maintenance of Chromosomes 5/6 (SMC5/6) complex as essential for cell viability when APOBEC3A is active. We observe an absence of APOBEC3A mutagenesis in human tumors with SMC5/6 dysfunction, consistent with synthetic lethality. Cancer cells depleted of SMC5/6 incur substantial genome damage from APOBEC3A activity during DNA replication. Further, APOBEC3A activity results in replication tract lengthening which is dependent on PrimPol, consistent with re-initiation of DNA synthesis downstream of APOBEC3A-induced lesions. Loss of SMC5/6 abrogates elongated replication tracts and increases DNA breaks upon APOBEC3A activity. Our findings indicate that replication fork lengthening reflects a DNA damage response to APOBEC3A activity that promotes genome stability in an SMC5/6-dependent manner. Therefore, SMC5/6 presents a potential therapeutic vulnerability in tumors with active APOBEC3A.

ATXN3 promotes proliferation, stemness and motility of clear cell renal cell carcinoma cells by regulating S100A8 ubiquitination.

Background: Clear cell renal cell carcinoma (ccRCC) is a dominant subtype of kidney cancer with a dismal outcome at advanced stages. Ataxin 3 (ATXN3) has been proven to play a cancer-promoting role in several tumors and is upregulated in the patients with renal cell carcinoma. Thus, the objective of this research is to examine the biological roles and underlying mechanisms of ATXN3 in ccRCC. Methods: Bioinformatics analysis was carried out to analyze ATXN3 expression in ccRCC tissues and patient survival. Gain- and loss-of-function assays were applied to explore the effect of ATXN3 on ccRCC cell malignant behavior in vitro. The effect of ATXN3 on the NF-κB pathway was assessed by Western blot and immunofluorescence staining. The binding between ATXN3 and S100A8 and the effect of ATXN3 on S100A8 ubiquitination were verified using coimmunoprecipitation. Results: ATXN3 was upregulated in ccRCC tissues and correlated with adverse patient outcome. ATXN3 overexpression facilitated the proliferation, stemness, invasion and migratory capacity of ccRCC cells, whereas silencing had the opposite effect. ATXN3 enhanced the activity of the NF-κB pathway. Silencing ATXN3 facilitated S100A8 ubiquitination. Rescue experiments demonstrated that S100A8 downregulation reversed the promoting effect of ATXN3 on malignant behavior and NF-κB pathway activation in ccRCC cells. Conclusion: ATXN3 exerts a cancer-promoting effect in ccRCC by regulating S100A8 ubiquitination. Therefore, targeting the ATXN3/S100A8/NF-κB axis may provide a novel underlying therapeutic strategy for ccRCC.

Bioorthogonal surface-enhanced Raman scattering flower-like nanoprobe with embedded standards for accurate cancer cell imaging.

Developing precise and effective strategies for cancer identification and imaging is attractive due to their importance for early cancer detection, prognosis, and subsequent treatment. Herein, we reported a novel bioorthogonal surface-enhanced Raman scattering (SERS) nanoprobe for accurate cancer cell imaging. A novel core-molecule-shell nanoflower (Au@4-MBN@Au) with rich electromagnetic hot spots and enhanced Raman scattering was first synthesized by optimizing the embedded concentrations of 4-mercaptobenzonitrile (4-MBN). Then, Au@4-MBN@Au was further modified with FA-PEG-SH molecules to acquire the bioorthogonal SERS nanoprobe Au@4-MBN@Au-PEG-FA. The SERS nanoprobe illustrated a robust and stable nitrile stretching vibration Raman signal (2223 cm-1) in the cellular silent region, ensuring high sensitivity and ultra-accuracy SERS imaging of cancer cells. Furthermore, cell imaging results demonstrated Au@4-MBN@Au-PEG-FA could recognize FR-positive HeLa cells with high selectivity due to the high affinity between folate receptor and folic acid. More notably, Au@4-MBN@Au-PEG-FA has been applied to identify FR-positive Hela cells from co-cultured cancer cells with similar morphology by SERS imaging for the first time. With improved signal-to-background ratio, high selectivity, and excellent stability, we anticipate the SERS nanoprobe Au@4-MBN@Au-PEG-FA could be applied for FR-related cancer theranostics and clinical detection in the future.