TRIB3-TRIM8 complex drives NAFLD progression by regulating HNF4α stability.

BACKGROUND & AIMS: Endoplasmic reticulum (ER) stress of hepatocytes plays a causative role in non-alcoholic fatty liver disease (NAFLD). Reduced expression of hepatic nuclear factor 4α (HNF4α) is a critical event in the pathogenesis of NAFLD and other liver diseases. Whether ER stress regulates HNF4α expression remains unknown. The aim of this study was to delineate the machinery of HNF4α protein degradation and explore a therapeutic strategy based on protecting HNF4α stability during NAFLD progression. METHODS: Correlation of HNF4α and tribbles homologue 3 (TRIB3), an ER stress sensor, was evaluated in human and mouse NAFLD tissues. RNA-sequencing, mass spectrometry analysis, co-immunoprecipitation, in vivo and in vitro ubiquitination assays were used to elucidate the mechanisms of TRIB3-mediated HNF4α degradation. Molecular docking and co-immunoprecipitation analyses were performed to identify a cell-penetrating peptide that ablates the TRIB3-HNF4α interaction. RESULTS: TRIB3 directly interacts with HNF4α and mediates ER stress-induced HNF4α degradation. TRIB3 recruits tripartite motif containing 8 (TRIM8) to form an E3 ligase complex that catalyzes K48-linked polyubiquitination of HNF4α on lysine 470. Abrogating the degradation of HNF4α attenuated the effect of TRIB3 on a diet-induced NAFLD model. Moreover, the TRIB3 gain-of-function variant p.Q84R is associated with NAFLD progression in patients, and induces lower HNF4α levels and more severe hepatic steatosis in mice. Importantly, disrupting the TRIB3-HNF4α interaction using a cell-penetrating peptide restores HNF4α levels and ameliorates NAFLD progression in mice. CONCLUSIONS: Our findings unravel the machinery of HNF4α protein degradation and indicate that targeting TRIB3-TRIM8 E3 complex-mediated HNF4α polyubiquitination may be an ideal strategy for NAFLD therapy. IMPACT AND IMPLICATIONS: Reduced expression of hepatic nuclear factor 4α (HNF4α) is a critical event in the pathogenesis of NAFLD and other liver diseases. However, the mechanism of HNF4α protein degradation remains unknown. Herein, we reveal that TRIB3-TRIM8 E3 ligase complex is responsible for HNF4α degradation during NAFLD. Inhibiting the TRIB3-HNF4α interaction effectively stabilized HNF4α protein levels and transcription factor activity in the liver and ameliorated TRIB3-mediated NAFLD progression. Our findings demonstrate that disturbing the TRIM8-TRIB3-HNF4α interaction may provide a novel approach to treat NAFLD and even other liver diseases by stabilizing the HNF4α protein.

An IL-10 homologue encoded by human cytomegalovirus is linked with the viral "footprint" in clinical samples.

Persistent infections with human cytomegalovirus (HCMV) affect the hosts' immune system and have been linked with chronic inflammation and cardiovascular disease. These effects may be influenced by a HCMV-encoded homologue of the anti-inflammatory cytokine, IL-10 (cmvIL-10). To assess this, we quantitated cmvIL-10 in plasma from renal transplant recipients (RTR) and healthy adults. Detectable levels of cmvIL-10 associated with seropositivity in RTR, but were found in some seronegative healthy adults. RTR with detectable cmvIL-10 had elevated interferon-γ T-cell responses to HCMV antigens, whilst cmvIL-10 in healthy adults associated with reduced populations of terminally-differentiated T-cells - a known "footprint" of HCMV. Plasma cmvIL-10 associated with lower VCAM-1 levels in healthy adults. The data suggest cmvIL-10 may suppress seroconversion and/or reduce the footprint of HCMV in healthy adults. This appears to be subverted in RTR by their high burden of HCMV and/or immune dysregulation associated with transplantation. A role for cmvIL-10 in protection of vascular health is discussed.

Real-world data on the efficacy and safety of hedgehog pathway inhibitors in patients with basal cell carcinoma: Experience of a tertiary Australian centre.

BACKGROUND: Basal cell carcinomas (BCCs) are the most common cancers worldwide. Although most BCCs are amenable to local treatment, there are limited therapeutic options for surgically unresectable locally advanced and metastatic BCCs. Activation of the sonic hedgehog signalling pathway plays a significant role in the development of most BCCs. Hedgehog pathway inhibitors (HPIs) can be used to inhibit this pathway. Efficacy and safety data on HPI use in Australia is scarce. OBJECTIVES: This study aims to present the effectiveness and safety of HPI at a tertiary dermatology referral centre. METHODS: We conducted a retrospective analysis of the clinical charts of all patients with BCC treated with an HPI at a tertiary Dermatology referral centre in New South Wales, Australia from 1 January 2016 to 1 July 2023. RESULTS: Twenty-three patients with BCCs were treated with an HPI; 11 locally advanced, 8 multiple, 3 basal cell naevus syndrome and 1 metastatic. All patients were of Caucasian background, with a median age of 56. Across 41 treatment cycles, the median treatment duration was 4 months. The overall response rate (ORR) was 20/23 (87%) and complete response (CR) rate was 9/23 (39%); patients treated with sonidegib achieved an ORR of 11/12 (92%) and CR of 4/12 (33%), and vismodegib-treated patients achieved an ORR of 9/11 (82%) and CR of 5/11 (45%). Patients who responded to HPI treatment also responded to a subsequent HPI rechallenge. Common treatment emergent adverse events (TEAEs) included muscle spasms, dysgeusia and alopecia. Dysgeusia was more frequent with vismodegib than sonidegib (p = 0.0001). There was no evidence to suggest a difference in other TEAEs between the two HPIs. Four treatment cycles were stopped due to grade 3 muscle spasm. CONCLUSIONS: In our cohort of 23 patients being treated with HPI, the ORR was 87% and CR was 39%. All patients who experienced TEAEs and had a drug holiday successfully responded to HPI rechallenge. TEAEs, particularly muscle spasms, are common reasons for treatment cessation. Clinicians should implement strategies to mitigate TEAE to improve drug survivability.

De Novo Linear Phosphorylation Site Motifs for BCR-ABL Kinase Revealed by Phospho-Proteomics in Yeast.

BCR-ABL is the oncogenic fusion product of tyrosine kinase ABL1 and a highly frequent driver of acute lymphocytic leukemia (ALL) and chronic myeloid leukemia (CML). The kinase activity of BCR-ABL is strongly elevated; however, changes of substrate specificity in comparison to wild-type ABL1 kinase are less well characterized. Here, we heterologously expressed full-length BCR-ABL kinases in yeast. We exploited the proteome of living yeast as an in vivo phospho-tyrosine substrate for assaying human kinase specificity. Phospho-proteomic analysis of ABL1 and BCR-ABL isoforms p190 and p210 yielded a high-confidence data set of 1127 phospho-tyrosine sites on 821 yeast proteins. We used this data set to generate linear phosphorylation site motifs for ABL1 and the oncogenic ABL1 fusion proteins. The oncogenic kinases yielded a substantially different linear motif when compared to ABL1. Kinase set enrichment analysis with human pY-sites that have high linear motif scores well-recalled BCR-ABL driven cancer cell lines from human phospho-proteome data sets.

New perspectives on the molecular mechanisms of stress signalling by the nucleotide guanosine tetraphosphate (ppGpp), an emerging regulator of photosynthesis in plants and algae.

The nucleotides guanosine tetraphosphate and guanosine pentaphosphate (together (p)ppGpp) are found in a wide range of prokaryotic and eukaryotic organisms where they are associated with stress signalling. In this review, we will discuss recent research highlighting the role of (p)ppGpp signalling as a conserved regulator of photosynthetic activity in the chloroplasts of plants and algae, and the latest discoveries that open up new perspectives on the emerging roles of (p)ppGpp in acclimation to environmental stress. We explore how rapid advances in the study of (p)ppGpp signalling in prokaryotes are now revealing large gaps in our understanding of the molecular mechanisms of signalling by (p)ppGpp and related nucleotides in plants and algae. Filling in these gaps is likely to lead to the discovery of conserved as well as new plant- and algal-specific (p)ppGpp signalling mechanisms that will offer new insights into the taming of the chloroplast and the regulation of stress tolerance.

Detrimental role of SIX1 in hepatic lipogenesis and fibrosis of non-alcoholic fatty liver disease.

BACKGROUND AND AIMS: Nonalcoholic fatty liver disease (NAFLD) is the most common liver disease worldwide. Aberrant lipid metabolism and accumulation of extracellular matrix proteins are hallmarks of the disease, but the underlying mechanisms are largely unknown. This study aims to elucidate the key role of sine oculis homeobox homologue 1 (SIX1) in the development of NAFLD. METHODS: Alb-Cre mice were administered the AAV9 vector for SIX1 liver-specific overexpression or knockdown. Metabolic disorders, hepatic steatosis, and inflammation were monitored in mice fed with HFHC or MCD diet. High throughput CUT&Tag analysis was employed to investigate the mechanism of SIX1 in diet-induced steatohepatitis. RESULTS: Here, we found increased SIX1 expression in the livers of NAFLD patients and animal models. Liver-specific overexpression of SIX1 using adeno-associated virus serotype 9 (AAV9) provoked more severe inflammation, metabolic disorders, and hepatic steatosis in the HFHC or MCD-induced mice model. Mechanistically, we demonstrated that SIX1 directly activated the expression of liver X receptor α (LXRα) and liver X receptor ß (LXRß), thus inducing de novo lipogenesis (DNL). In addition, our results also illustrated a critical role of SIX1 in regulating the TGF-ß pathway by increasing the levels of type I and II TGF-ß receptor (TGFßRI/TGFßRII) in hepatic stellate cells (HSCs). Finally, we found that liver-specific SIX1 deficiency could ameliorate diet-induced NAFLD pathogenesis. CONCLUSION: Our findings suggest a detrimental function of SIX1 in the progression of NAFLD. The direct regulation of LXRα/ß and TGF-ß signalling by SIX1 provides a new regulatory mechanism in hepatic steatosis and fibrosis.

Genetic compensation response could exist in colorectal cancer: UPF3A upregulates the oncogenic homologue gene SRSF3 expression corresponding to SRSF6 to promote colorectal cancer metastasis.

BACKGROUND: Genetic compensation response (GCR) is a mechanism that maintains the robustness of functional genes, which has been recently identified. Whether GCR exists in tumors and its effects on tumor progression remains unknown. METHODS: Whole exome sequencing was performed to identify premature termination codon (PTC) gene mutations in colorectal cancer (CRC) tissues. RNA sequencing, Cancer Cell Line Encyclopedia database analysis, and high-throughput output of homologous genes using the Ensemble genome database were performed to further identify homologous genes of target PTC gene mutations. RESULTS: Serine and arginine-rich splicing factor 3 (SRSF3) increased the invasion ability in CRC cells and could be the target gene of up-frameshift 3A (UPF3A). The deletion of the 660th base A in the coding sequence region of SRSF6 caused a frameshift mutation of serine at position 220 (s220fs), which contributed to a PTC UAA termination of translation in HCT116 cells. We further found that SRSF3 was the only homologue of SRSF6 with a frameshift mutation. The transfection of s220fs of SRSF6 into HCT116 cells led to upregulation of its corresponding oncogenic homologue gene SRSF3 expression to promote CRC metastasis. SRSF3 was highly expressed in CRC liver metastases and was positively correlated with UPF3A expression and contributed to poor prognosis. CONCLUSION: GCR may exist in CRC and exert effects on the progression of CRC. Targeted inhibition of UPF3A could reduce the GCR effects and suppress the expression of oncogenic homologue genes corresponding to PTC mutations, indicating a novel therapeutic strategy for treatment of CRC metastasis.

Overexpression of SMAD7 improves the function of EGFR-targeted human CAR-T cells against non-small-cell lung cancer.

BACKGROUND AND OBJECTIVE: Recent advancements in immunotherapy led to the development of Chimeric antigen receptor (CAR) T-cell therapy. CAR-T cell therapy in non-small cell lung cancer (NSCLC) is hindered by overexpression of transforming growth factor (TGFß) in the cancer cells that have a negative regulatory role on T-cells activity. This study characterized CAR-T with overexpression of mothers against decapentaplegic homologue 7 (SMAD), a negative regulator of TGFß downstream signalling. METHODS: We have generated three types of CAR-T: epidermal growth factor receptor (EGFR)-CAR-T, EGFR-dominant-negative TGFbeta receptor 2 (DNR)-CAR-T, and EGFR-SMAD7-CAR-T by transducing human T-cells with the lentivirus constructs. We characterized the proliferation, expression of proinflammatory cytokines, activation profile, and lysis capacity in co-cultures with A549 lung carcinoma cells with and without TGFß neutralizing antibodies. We also tested the therapeutic potential of EGFR-SMAD7-CAR-T in the A549 cells tumour-bearing mice model. RESULTS: Both EGFR-DNR-CAR-T and EGFR-SMAD7-CAR-T demonstrated a higher proliferation rate and lysis capacity to A549 than traditional EGFR-CAR-T. Neutralization of TGFß by the antibodies resulted in increased performance of EGFR-CAR-T. In vivo, both EGFR-DNR-CAR-T and EGFR-SMAD7-CAR-T resulted in complete tumour resorption by day 20, whereas conventional CAR-T only has a partial effect. CONCLUSION: We demonstrated the high efficacy and resistance to negative TGFß regulation of EGFR-SMAD7-CAR-T comparable with EGFR-DNR-CAR-T and without the systemic effect of TGFß inhibition.

Analysis of CRB1 Pathogenic Variants Correctable with CRISPR Base and Prime Editing.

The mouse and human retina contain three major Crumbs homologue-1 (CRB1) isoforms. CRB1-A and CRB1-B have cell-type-specific expression patterns making the choice of gene augmentation strategy unclear. Gene editing may be a viable alternative for the amelioration of CRB1-associated retinal degenerations. To assess the prevalence and spectrum of CRB1-associated pathogenic variants amenable to base and prime editing, we carried out an analysis of the Leiden Open Variation Database. Editable variants accounted for 54.5% for base editing and 99.8% for prime editing of all CRB1 pathogenic variants in the Leiden Open Variation Database. The 10 most common editable pathogenic variants for CRB1 accounted for 34.95% of all pathogenic variants, with the c.2843G>A, p.(Cys948Tyr) being the most common editable CRB1 variant. These findings outline the next step toward developing base and prime editing therapeutics as an alternative to gene augmentation for the amelioration of CRB1-associated retinal degenerations.

Generation of CRB1 RP Patient-Derived iPSCs and a CRISPR/Cas9-Mediated Homology-Directed Repair Strategy for the CRB1 c.2480G>T Mutation.

Mutations in the Crumbs-homologue-1 (CRB1) gene lead to a spectrum of severe inherited retinal diseases, including retinitis pigmentosa (RP). The establishment of a genotype-phenotype correlation in CRB1 patients has been difficult due to the substantial variability and phenotypic overlap between CRB1-associated diseases. This phenotypic modulation may be due to several factors, including genetic modifiers, deep intronic mutations, isoform diversity, and copy number variations. Induced pluripotent stem cell (iPSC)-derived patient retinal organoids are novel tools that can provide sensitive, quantitative, and scalable phenotypic assays. CRB1 RP patient iPSC-derived retinal organoids have shown reproducible phenotypes compared to healthy retinal organoids. However, having genetically defined iPSC isogenic controls that take into account potential phenotypic modulation is crucial. In this study, we generated iPSC from an early-onset CRB1 patient and developed a correction strategy for the c.2480G>T, p.(Gly827Val) CRB1 mutation using CRISPR/Cas9-mediated homology-directed repair.

Crystal Structure of the Catalytic Domain of a Botulinum Neurotoxin Homologue from Enterococcus faecium: Potential Insights into Substrate Recognition.

Clostridium botulinum neurotoxins (BoNTs) are the most potent toxins known, causing the deadly disease botulism. They function through Zn2+-dependent endopeptidase cleavage of SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins, preventing vesicular fusion and subsequent neurotransmitter release from motor neurons. Several serotypes of BoNTs produced by Clostridium botulinum (BoNT/A-/G and/X) have been well-characterised over the years. However, a BoNT-like gene (homologue of BoNT) was recently identified in the non-clostridial species, Enterococcus faecium, which is the leading cause of hospital-acquired multi-drug resistant infections. Here, we report the crystal structure of the catalytic domain of a BoNT homologue from Enterococcus faecium (LC/En) at 2.0 Å resolution. Detailed structural analysis in comparison with the full-length BoNT/En AlphaFold2-predicted structure, LC/A (from BoNT/A), and LC/F (from BoNT/F) revealed putative subsites and exosites (including loops 1-5) involved in recognition of LC/En substrates. LC/En also appears to possess a conserved autoproteolytic cleavage site whose function is yet to be established.

[Significance of IgA isotype of anti-v-raf murine sarcoma viral oncogene homologue B1 antibody in rheumatoid arthritis].

OBJECTIVE: To detect serum IgA isotype of anti-v-raf murine sarcoma viral oncogene homologue B1 (BRAF) antibody levels in the rheumatoid arthritis (RA) patients in order to investigate their clinical significance in RA. METHODS: Serum samples were obtained from 61 RA patients, 21 osteoarthritis (OA) patients, 16 systemic lupus erythematosus (SLE) patients, 16 gout patients, 16 Sjögren's syndrome (SS) patients and 22 healthy controls. IgA isotype of anti-BRAF antibody levels in the sera were examined by enzyme-linked immunosorbent assay (ELISA). The associations between IgA isotype of anti-BRAF antibody levels and the clinical features including age, disease duration and laboratory parameters including erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), rheumatoid factor (RF), disease activity score in 28 joints (DAS28), anti-cyclic citrullinated peptide (anti-CCP) antibody, immunoglobulin and BRAF protein levels in the RA patients were evaluated. Data analyses were performed by using SPSS 19.0 program. RESULTS: The serum IgA isotype of anti-BRAF antibody levels in the RA patients were significantly higher than in the SLE, gout, OA patients and healthy controls, the P value was 0.011, < 0.001, < 0.001 and < 0.001, respectively. The serum IgA isotype of anti-BRAF antibody levels in the SS patients were also significantly higher than in the SLE, gout, OA patients and healthy controls, the P value was 0.029, 0.004, 0.001 and < 0.001, respectively. However, there was no difference between the RA and SS patients (P=0.762). IgA isotype of anti-BRAF antibody was measurable in the RA patients without RF, anti-CCP antibody or anti-keratin antibody (AKA) antibodies. The levels of IgA isotype of anti-BRAF antibody in the RA patients did not show any correlation with clinical features such as age and disease duration or laboratory parameters including ESR, CRP, RF, DAS28, anti-CCP antibody, immunoglobulin and BRAF protein levels. Compared with the clinical features and laboratory indexes of normal and elevated levels of IgA isotype of anti-BRAF antibody groups in the RA patients, there was no significant differences between the two groups in age, disease duration, ESR, CRP, RF, DAS28, anti-CCP antibody, immunoglobulin or BRAF protein levels. CONCLUSION: The elevated level of IgA isotype of anti-BRAF antibody in the RA patients showed that IgA isotype of anti-BRAF antibody might play a role in the pathogenesis of RA. Furthermore, detection of IgA isotype of anti-BRAF antibody in the serum negative RA patients showed that it might be helpful for the diagnosis of the serum negative RA patients.

Skeletal muscle proteome expression differentiates severity of cancer cachexia in mice and identifies loss of fragile X mental retardation syndrome-related protein 1.

Tandem mass tag (TMT)-based quantitative proteomics was used to examine protein expression in skeletal muscle from mice with moderate and severe cancer cachexia to study mechanisms underlying varied cachexia severity. Weight loss of 10% (moderate) and 20% (severe) was induced by injection of colon-26 cancer cells in 10-week old Balb/c mice. In moderate cachexia, enriched pathways reflected fibrin formation, integrin/mitogen-activated protein kinase (MAPK) signaling, and innate immune system, suggesting an acute phase response and fibrosis. These pathways remained enriched in severe cachexia; however, energy-yielding pathways housed in mitochondria were prominent additions to the severe state. These enrichments suggest distinct muscle proteome expression patterns that differentiate cachexia severity. When analyzed with two other mouse models, eight differentially expressed targets were shared including serine protease inhibitor A3N (Serpina3n), synaptophysin-like protein 2 (Sypl2), Isocitrate dehydrogenase [NAD] subunit alpha, mitochondrial (Idh3a), peroxisomal acyl-coenzyme A oxidase 1 (Acox1), collagen alpha-1(VI) chain (Col6a1), myozenin 3 (Myoz3), UDP-glucose pyrophosphorylase (Ugp2), and solute carrier family 41 member 3 (Slc41a3). Acox1 and Idh3a control lipid oxidation and NADH generation in the TCA cycle, respectively, and Col6a1 comprises part of type VI collagen with reported profibrotic functions, suggesting influential roles in cachexia. A potential target was identified in fragile X mental retardation syndrome-related protein 1 (FXR1), an RNA-binding protein not previously implicated in cancer cachexia. FXR1 decreased in cachexia and related linearly with weight change and myofiber size. These findings suggest distinct mechanisms associated with cachexia severity and potential biomarkers and therapeutic targets.

Balance between bacterial extracellular matrix production and intramacrophage proliferation by a Salmonella-specific SPI-2-encoded transcription factor.

Biosynthesis and secretion of a complex extracellular matrix (EM) is a hallmark of Salmonella biofilm formation, impacting on its relationship with both the environment and the host. Cellulose is a major component of Salmonella EM. It is considered an anti-virulence factor because it interferes with Salmonella proliferation inside macrophages and virulence in mice. Its synthesis is stimulated by CsgD, the master regulator of biofilm formation in enterobacteria, which in turn is under the control of MlrA, a MerR-like transcription factor. In this work, we identified a SPI-2-encoded Salmonella-specific transcription factor homolog to MlrA, MlrB, that represses transcription of its downstream gene, orf319, and of csgD inside host cells. MlrB is induced in laboratory media mimicking intracellular conditions and inside macrophages, and it is required for intramacrophage proliferation. An increased csgD expression is observed in the absence of MlrB inside host cells. Interestingly, inactivation of the CsgD-controlled cellulose synthase-coding gene restored intramacrophage proliferation to rates comparable to wild-type bacteria in the absence of MlrB. These data indicate that MlrB represses CsgD expression inside host cells and suggest that this repression lowers the activation of the cellulose synthase. Our findings provide a novel link between biofilm formation and Salmonella virulence.

The anterior gradient homologue 2 (AGR2) co-localises with the glucose-regulated protein 78 (GRP78) in cancer stem cells, and is critical for the survival and drug resistance of recurrent glioblastoma: in situ and in vitro analyses.

BACKGROUND: Glioblastomas (GBs) are characterised as one of the most aggressive primary central nervous system tumours (CNSTs). Single-cell sequencing analysis identified the presence of a highly heterogeneous population of cancer stem cells (CSCs). The proteins anterior gradient homologue 2 (AGR2) and glucose-regulated protein 78 (GRP78) are known to play critical roles in regulating unfolded protein response (UPR) machinery. The UPR machinery influences cell survival, migration, invasion and drug resistance. Hence, we investigated the role of AGR2 in drug-resistant recurrent glioblastoma cells. METHODS: Immunofluorescence, biological assessments and whole exome sequencing analyses were completed under in situ and in vitro conditions. Cells were treated with CNSTs clinical/preclinical drugs taxol, cisplatin, irinotecan, MCK8866, etoposide, and temozolomide, then resistant cells were analysed for the expression of AGR2. AGR2 was repressed using single and double siRNA transfections and combined with either temozolomide or irinotecan. RESULTS: Genomic and biological characterisations of the AGR2-expressed Jed66_GB and Jed41_GB recurrent glioblastoma tissues and cell lines showed features consistent with glioblastoma. Immunofluorescence data indicated that AGR2 co-localised with the UPR marker GRP78 in both the tissue and their corresponding primary cell lines. AGR2 and GRP78 were highly expressed in glioblastoma CSCs. Following treatment with the aforementioned drugs, all drug-surviving cells showed high expression of AGR2. Prolonged siRNA repression of a particular region in AGR2 exon 2 reduced AGR2 protein expression and led to lower cell densities in both cell lines. Co-treatments using AGR2 exon 2B siRNA in conjunction with temozolomide or irinotecan had partially synergistic effects. The slight reduction of AGR2 expression increased nuclear Caspase-3 activation in both cell lines and caused multinucleation in the Jed66_GB cell line. CONCLUSIONS: AGR2 is highly expressed in UPR-active CSCs and drug-resistant GB cells, and its repression leads to apoptosis, via multiple pathways.

Global Historical Production, Use, In-Use Stocks, and Emissions of Short-, Medium-, and Long-Chain Chlorinated Paraffins.

The last few decades have seen ubiquitous and increasing contamination of chlorinated paraffins (CPs) worldwide. Here, we develop the first global inventories of production, use, in-use stocks, and emissions of total CPs, including the short-, medium- and long-chain components (SCCPs, MCCPs, and LCCPs) during 1930-2020 using a dynamic substance flow analysis model named Chemical in Products Comprehensive Anthroposhpheric Fate Estimation. The model estimates that a total of ∼33 million metric tons of CPs have been produced and used globally, ∼40% of which still resided in in-use products by 2020 and is available for long-term emissions in the next decades. Global cumulative emissions of CPs have increased to ∼5.2 million metric tons by 2020, with SCCPs, MCCPs, and LCCPs accounting for ∼30, 40, and 30%, respectively. While the production, use, and emissions of CPs started declining in regions such as Western Europe, they remain high in China. The model also suggests that homologues with 10, 14, and 22-23 carbons were predominant in the cumulatively produced and emitted SCCPs, MCCPs, and LCCPs, respectively. The emission estimates were evaluated by generating environmental concentrations that are comparable to literature-reported environmental monitoring data. Our estimates provide opportunities to link the environmental fate and occurrence of CPs to emission sources and lay the basis for future risk-reduction strategies of CPs around the world.

Influence of elongation and desaturation on chemosensory properties in acrylates and their corresponding 1-alken-3-ones.

Acrylates as well as 1-alken-3-ones are both known to be odour active substances but are generally identified in different materials. Nonetheless, butyl acrylate and 1-octen-3-one were both found to elicit a similar mushroom-like odour in previous studies. This led to the question of whether acrylates and enones with the same overall chain length generally elicited similar odours and whether they had similar odour thresholds. Overall, most of the investigated substances showed a mushroom-like, geranium-like or fruity odour. In contrast, short chained substances elicited garlic-like, lighter gas-like or glue-like, odour qualities, suggesting a correlation between the odour quality and the overall chain length. The results showed that only between the analogue structures butyl acrylate and 1-octen-3-one as well as hexyl acrylate and 1-decen-3-one could similar odour qualities be observed. All investigated substances showed low odour threshold values in air between 0.0032 ng/lair (1-hexen-3-one) and 55 ng/lair (1-dodecen-3-one). Overall, 1-alken-3-ones revealed a higher dependency on the chain length when compared to their respective acrylates. The introduction of a second terminal double bond led to a decrease of OT values in case of the acrylates and to an increase in case of the ketones that neither contained a second terminal double bond nor a double bond located close to the carbonyl group. Despite their structural similarities, the results suggest that both substance classes are perceived in a different manner and are therefore likely to be recognized by different types of receptors or are related to different activation patterns in multi-receptor stimulation processes.

EZH2 promotes the progression of osteosarcoma through the activation of the AKT/GSK3ß pathway.

Enhancer of zeste homologue 2 (EZH2) is a clarified promoter in a list of tumours, including osteosarcoma (OS). Our research was projected to define the mechanism involved in EZH2-mediated OS progression through the protein kinase B (AKT)/glycogen synthase kinase 3ß (GSK3ß) pathway. EZH2 expression was tested in 66 OS tissues and five osteosarcoma cell lines (143B, SJSA-1, HOS, MG63, and U2OS). In HOS and U2OS cells, cellular malignant characteristics, and the markers of the AKT/GSK3ß signalling pathway were measured when EZH2 was silenced or overexpressed. Meanwhile, rescue assays were implemented to observe whether the AKT/GSK3ß signalling pathway inhibitor (MK-2206) could affect the role of overexpressed EZH2 in OS cells. EZH2 was up-regulated in tumour tissues of OS patients. OS cell lines (HOS and U2OS) showed impairments of proliferative, migratory, invasive and anti-apoptotic properties when EZH2 was silenced. Downregulated EZH2 inhibited the activation of the AKT/GSK3 signalling pathway. However, the situation in HOS and U2OS cells over-expressing EZH2 was opposite. MK-2206 erased EZH2 up-regulation-induced promotion of OS cell growth. It is demonstrated that EZH2 promotes the progression of OS via inducing the activation of the AKT/GSK3ß pathway, offering a therapeutic direction for OS treatment.

Impact of a Novel PagR-like Transcriptional Regulator on Cereulide Toxin Synthesis in Emetic Bacillus cereus.

The emetic type of foodborne disease caused by Bacillus cereus is produced by the small peptide toxin cereulide. The genetic locus encoding the Ces nonribosomal peptide synthetase (CesNRPS) multienzyme machinery is located on a 270 kb megaplasmid, designated pCER270, which shares its backbone with the Bacillus anthracis toxin plasmid pXO1. Although the ces genes are plasmid-borne, the chromosomally encoded pleiotropic transcriptional factors CodY and AbrB are key players in the control of ces transcription. Since these proteins only repress cereulide synthesis during earlier growth phases, other factors must be involved in the strict control of ces expression and its embedment in the bacterial life cycle. In silico genome analysis revealed that pCER270 carries a putative ArsR/SmtB family transcription factor showing high homology to PagR from B. anthracis. As PagR plays a crucial role in the regulation of the protective antigen gene pagA, which forms part of anthrax toxin, we used a gene-inactivation approach, combined with electrophoretic mobility shift assays and a bacterial two-hybrid system for dissecting the role of the PagR homologue PagRBc in the regulation of cereulide synthesis. Our results highlight that the plasmid-encoded transcriptional regulator PagRBc plays an important role in the complex and multilayered process of cereulide synthesis.

The SCO2102 Protein Harbouring a DnaA II Protein-Interaction Domain Is Essential for the SCO2103 Methylenetetrahydrofolate Reductase Positioning at Streptomyces Sporulating Hyphae, Enhancing DNA Replication during Sporulation.

Streptomyces DNA replication starts with the DnaA binding to the origin of replication. Differently to most bacteria, cytokinesis only occurs during sporulation. Cytokinesis is modulated by the divisome, an orderly succession of proteins initiated by FtsZ. Here, we characterised SCO2102, a protein harbouring a DnaA II protein-protein interaction domain highly conserved in Streptomyces. The ΔSCO2102 knockout shows highly delayed sporulation. SCO2102-mCherry frequently co-localises with FtsZ-eGFP during sporulation and greatly reduces FtsZ-eGFP Z-ladder formation, suggesting a role of SCO2102 in sporulation. SCO2102 localises up-stream of SCO2103, a methylenetetrahydrofolate reductase involved in methionine and dTMP synthesis. SCO2102/SCO2103 expression is highly regulated, involving two promoters and a conditional transcription terminator. The ΔSCO2103 knockout shows reduced DNA synthesis and a non-sporulating phenotype. SCO2102-mCherry co-localises with SCO2103-eGFP during sporulation, and SCO2102 is essential for the SCO2103 positioning at sporulating hyphae, since SCO2103-eGFP fluorescent spots are absent in the ΔSCO2102 knockout. We propose a model in which SCO2102 positions SCO2103 in sporulating hyphae, facilitating nucleotide biosynthesis for chromosomal replication. To the best of our knowledge, SCO2102 is the first protein harbouring a DnaA II domain specifically found during sporulation, whereas SCO2103 is the first methylenetetrahydrofolate reductase found to be essential for Streptomyces sporulation.