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OBJECTIVE: To explore susceptibility loci associated with uveitis in Behçet's disease (BD).METHODS: We conducted a 2-stage study, consisting of a genome-wide association study (GWAS) stage and a replication stage, in a Chinese population. The GWAS stage included 978 cases with BD-related uveitis and 4,388 controls, and the replication stage included 953 cases with BD-related uveitis and 2,129 controls. Luciferase reporter analysis and chromatin immunoprecipitation assay were performed to explore the functional role of susceptibility genetic variants near ZMIZ1.RESULTS: Three independent HLA alleles (HLA-B51 [3.75 × 10-190 ], HLA-A26 [1.50 × 10-18 ], and HLA-C0704 [3.44 × 10-16 ]) were identified as having a genome-wide association with BD-related uveitis. In the non-HLA region, in addition to confirming 7 previously reported loci, we identified 22 novel susceptibility variants located in 16 loci. Meta-analysis of the Chinese cohort consisting of 1,931 cases and 6,517 controls and a published Japanese cohort of 611 cases and 737 controls showed genome-wide significant associations with ZMIZ1, RPS6KA4, IL10RA, SIPA1-FIBP-FOSL1, and VAMP1. Functional experiments demonstrated that genetic variants of ZMIZ1 were associated with enhanced transcription activity and increased expression of ZMIZ1.CONCLUSION: This GWAS study identified a novel set of genetic variants that are associated with susceptibility to uveitis in BD. These findings enrich our understanding of the contribution of genetic factors to the disease.

The circRNAs, a new subclass of non-coding RNAs that are catalyzed by RNA-binding proteins (RBPs), have been reported to be associated with the progression of multiple types of cancer. We previously discovered that heterogeneous nuclear ribonucleoprotein L (HnRNP-L), a multi-functional RBP, is associated with pro-proliferation and anti-apoptosis activities in prostate tumor cells. In this study, we aim to establish the biological relevance of circCSPP1 (a newly discovered signature circRNA in prostate cancer [PCa]) and HnRNP-L to prostate cancer progression. First, we demonstrated that circCSPP1 expression was higher in prostate cancer tissues than in benign tissues and higher in prostate cancer cells than in benign cells. Then, the in vitro gain- and loss-of-function experiments showed that the circCSPP1 expression in prostate cancer cells was regulated by HnRNP-L, and the increased circCSPP1 significantly induced autophagy, which led to an enhanced potential in proliferation, migration, and invasion of prostate cancer cells. These results were consistent with the in vivo experiment where increased or decreased circCSPP1 was associated with higher or slower growth rate in grafted tumors. Finally, we demonstrated the potential competing endogenous RNA network, involving circCSPP1, miR-520h, and early growth response factor 1 (EGR1), in prostate cancer cells, which may play an important role in prostate cancer progression. Our study indicated that the increase in circCSPP1 in prostate cancer, which may be catalyzed by HnRNP-L, can induce cellular autophagy through the circCSPP1-miR-520h-EGR1 axis, leading to the progression of prostate tumor. This newly discovered circRNA biomarker may be used for clinical prognosis of prostate cancer as well as for development of novel therapy plans.

Abnormal activation of the mechanistic target of rapamycin (mTOR) signaling is commonly observed in many cancers and attracts extensive attention as an oncology drug discovery target, which is encouraged by the success of rapamycin and its analogs (rapalogs) in treatment of mTORC1-hyperactive cancers in both pre-clinic models and clinical trials. However, rapamycin and existing rapalogs have typically short-lasting partial responses due to drug resistance, thereby triggering our interest to investigate a potential mTORC1 inhibitor that is mechanistically different from rapamycin. Here, we report that hayatine, a derivative from Cissampelos, can serve as a potential mTORC1 inhibitor selected from a natural compound library. The unique properties owned by hayatine such as downregulation of mTORC1 activities, induction of mTORC1's translocation to lysosomes followed by autophagy, and suppression on cancer cell growth, strongly emphasize its role as a potential mTORC1 inhibitor. Mechanistically, we found that hayatine disrupts the interaction between mTORC1 complex and its lysosomal adaptor RagA/C by binding to the hydrophobic loop of RagC, leading to mTORC1 inhibition that holds great promise to overcome rapamycin resistance. Taken together, our data shed light on an innovative strategy using structural interruption-based mTORC1 inhibitors for cancer treatment.

The ability to control translation of endogenous or exogenous RNAs in eukaryotic cells would facilitate a variety of biotechnological applications. Current strategies are limited by low fold changes in transgene output and the size of trigger RNAs (trRNAs). Here we introduce eukaryotic toehold switches (eToeholds) as modular riboregulators. eToeholds contain internal ribosome entry site sequences and form inhibitory loops in the absence of a specific trRNA. When the trRNA is present, eToeholds anneal to it, disrupting the inhibitory loops and allowing translation. Through optimization of RNA annealing, we achieved up to 16-fold induction of transgene expression in mammalian cells. We demonstrate that eToeholds can discriminate among viral infection status, presence or absence of gene expression and cell types based on the presence of exogenous or endogenous RNA transcripts.

Bromodomain and extra-terminal domain (BET) family proteins are epigenetic readers that play a critical role in oncogenesis by controlling the expression of oncogenes such as c-Myc. Targeting BET family proteins has recently emerged as a promising anticancer strategy. However, the molecular mechanisms by which cancer cells respond to BET inhibition are not well understood. In this study, we found that inducing the degradation of BET proteins by the proteolysis targeting chimeras (PROTAC) approach potently suppressed the growth of colorectal cancer (CRC) including patient-derived tumors. Mechanistically, BET degradation transcriptionally activates Death Receptor 5 (DR5) to trigger immunogenic cell death (ICD) in CRC cells. Enhanced DR5 induction further sensitizes CRC cells with a mutation in Speckle-type POZ protein (SPOP). Furthermore, DR5 is indispensable for a striking antitumor effect of combining BET degradation and anti-PD-1 antibody, which was well tolerated in mice and almost eradicated syngeneic tumors. Our results demonstrate that BET degradation triggers DR5-mediated ICD to potently suppress CRC and potentiate immune checkpoint blockade. These results provide a rationale, mechanistic insights, and potential biomarkers for developing a precision CRC therapy by inducing BET protein degradation.

Ferroptosis is a lipid peroxidation-dependent cell death caused by metabolic dysfunction. Ferroptosis-associated enzymes are promising therapeutic targets for cancer treatment. However, such therapeutic strategies show limited efficacy due to drug resistance and other largely unknown underlying mechanisms. Here we report that cystine transporter SLC7A11 is upregulated in lung cancer stem-like cells (CSLC) and can be activated by stem cell transcriptional factor SOX2. Mutation of SOX2 binding site in SLC7A11 promoter reduced SLC7A11 expression and increased sensitivity to ferroptosis in cancer cells. Oxidation at Cys265 of SOX2 inhibited its activity and decreased the self-renewal capacity of CSLCs. Moreover, tumors with high SOX2 expression were more resistant to ferroptosis, and SLC7A11 expression was positively correlated with SOX2 in both mouse and human lung cancer tissue. Together, our study provides a mechanism by which cancer cells evade ferroptosis and suggests that oxidation of SOX2 can be a potential therapeutic target for cancer treatment. Significance: This study uncovers a SOX2–SLC7A11 regulatory axis that confers resistance to ferroptosis in lung cancer stem-like cells.

Understanding intratumoral heterogeneity-the molecular variation among cells within a tumor-promises to address outstanding questions in cancer biology and improve the diagnosis and treatment of specific cancer subtypes. Single-cell analyses, especially RNA sequencing and other genomics modalities, have been transformative in revealing novel biomarkers and molecular regulators associated with tumor growth, metastasis and drug resistance. However, these approaches fail to provide a complete picture of tumor biology, as information on cellular location within the tumor microenvironment is lost. New technologies leveraging multiplexed fluorescence, DNA, RNA and isotope labeling enable the detection of tens to thousands of cancer subclones or molecular biomarkers within their native spatial context. The expeditious growth in these techniques, along with methods for multiomics data integration, promises to yield a more comprehensive understanding of cell-to-cell variation within and between individual tumors. Here we provide the current state and future perspectives on the spatial technologies expected to drive the next generation of research and diagnostic and therapeutic strategies for cancer.

The Middle Jurassic flora from Baiyanghe (Baiyang River) area in Junggar Basin, Xinjiang, China, was first reported by Seward (1911) with fossils collected mainly at Ak-djar (Baiyanghe) in Diam (Baiyang) River valley by Russian geologist V.A. Obruchev in 1904–1909. The recent new study of this flora has advanced the taxonomy, particularly for its ferns, including Equisetites, Neocalamites, Coniopteris and Raphaelia. The achievement is related to the Sino-German co-research team working in this basin since 1997, led by V. Mosbrugger and G. Sun.

Programmed cell death‐ligand 1 (PD‐L1) expressed on cancer cells can cause immune escape of non‐small‐cell lung cancer (NSCLC). Elucidation of the regulatory mechanisms of the PD‐L1 expression is a prerequisite for establishing new tumor immunotherapy strategies. Ubiquitin C‐terminal hydrolase L1 (UCHL1) is a regulator of cellular signaling transduction that is aberrantly expressed in NSCLC. However, it is not known whether UCHL1 regulates the expression of PD‐L1 in NSCLC cells. In the present study, we found that UCHL1 promotes the expression of PD‐L1 in NSCLC cell lines. In addition, UCHL1 expressed in NSCLC cells inhibited activation of Jurkat cells through upregulation of PD‐L1 expression in in vitro experiments. Moreover, UCHL1 upregulates PD‐L1 expression through facilitating activation of the AKT‐P65 signaling pathway. In conclusion, these results indicated that UCHL1 promoted PD‐L1 expression in NSCLC cells. This finding implied that inhibition of UCHL1 might suppress immune escape of NSCLC through downregulation of PD‐L1 expression in NSCLC cells., Deubiquitinase ubiquitin C‐terminal hydrolase L1 (UCHL1) is a regulator of cellular signaling transduction that is aberrantly expressed in non‐small‐cell lung cancer (NSCLC). We analyzed the relationship between deubiquitinase UCHL1 and immune checkpoint molecule programmed cell death‐ligand 1 (PD‐L1) in NSCLC cells. Our results showed that UCHL1 promoted expression of PD‐L1 through the Akt‐P65 signaling axis.

Exploring and utilizing the agronomic potential of deep-underground is one of the ways to cope with the challenges of sudden environmental change on agriculture. Understanding the effects of environmental stresses on the morphological and physiological indicators of crop seeds after their storage deep-underground is crucial to developing and implementing strategies for agriculture in the deep-underground space. In this study, we stored canola seeds in envelopes or sealed packages at 0, 240, 690, and 1,410 m in a gold mine. Seeds in envelopes were retrieved at 42, 66, 90, and 227 days of storage, whereas seeds in sealed packages were retrieved at 66 and 227 days of storage. The germination tests were conducted to investigate the effects of storage depth, duration, and packing method on stored and non-stored seeds. Results showed that increased depth and duration led to decreased seed germination rate, with the germination and vigor indexes also descending to varying degrees. Increased hypocotyl length and biomass accumulation suggested that deep-underground environment had more significant compensatory effect on seed germination. For all indicators, the performance of seeds sealed in packages was superior to those stored in envelopes. Regression analysis showed that it was difficult to obtain the optimal value of each indicator simultaneously. The successful germination experiment foreshadowed the possibilities of deep-underground agriculture in the future.

Aims: The genetic association between Behcet's disease susceptibility and IL-10 has been confirmed in multiple cohorts, but the underlying mechanism of this association remains unclear. Materials & methods: We combined public resources and laboratory experiments (electrophoretic mobility shift assays, chromatin immunoprecipitation, luciferase reporter gene and CRISPR/Cas9 genome editing) to analyze transcription factor binding and enhancer activity controlling IL-10 expression. Results & conclusion: The T allele of noncoding rs3024490 within super-enhancer elements is able to specifically bind TBX1 and, in turn, promotes the enhancer activity and increased expression of IL-10. However, a relative deficiency in TBX1 in Behcet's disease patients leads to the low expression of IL-10 and increased risk of developing Behcet's disease.Lay abstract Behcet's disease is an important cause of blindness. Previous reports show that genetic factors are linked with this disease, although the exact genetic mechanism is unclear. Many of these genetic factors are involved in the control of the immune response, including a large family of proteins known as cytokines. Some of the cytokines are proinflammatory while others can dampen the inflammatory response. An example of the latter is the IL-10 cytokine. We found that individuals carrying a specific site in the noncoding region of the IL-10 gene had a higher risk of Behcet's disease than other noncarriers. This study was designed to further investigate the biological mechanisms explaining the role of the specific site in the development of Behcet's disease. The results show that this specific site affects the binding of an important transcription factor, TBX1, which reduces IL-10 production. The dysregulated control of IL-10 explains why individuals with this genetic trait are more susceptible to developing Behcet's disease.

Alpine musk deer, Moschus chrysogaster, a solitary, primitive ungulate inhabiting high elevation areas (3000–4500 m) is an endangered species facing threat of extinction globally due to excessive h...

In lakes, suspended inorganic particles and dissolved substance are able to absorb or scatter different light wavelengths, leading to the changes of underwater light spectra which are highly related to the water quality. In turn, such changes could form environmental filtering for phytoplankton community to select particular algal populations via intensive competition for light resources. As an example, eutrophic lakes where underwater light spectra changed dramatically have a result of cyanobacterial blooms. In this study, in order to test the effect of light spectrum on growth and competition of green algae and cyanobacteria, Chlorella pyrenoidosa (a common green alga) and Microcystis aeruginosa (a bloom-forming cyanobacterium) grew and competed under three light colors: white (400–700 nm), red (620–700 nm), and blue (410–490 nm) light. Mono- and co-cultured systems were designed and population dynamics of the two species were monitored. The Lotka-Volterra model was used to quantify interspecific competition. Moreover, their photosynthetic activities were measured in mono-cultures. Results showed that in mono-cultures, red light was more favorable for M. aeruginosa, while blue light promoted the growth of C. pyrenoidosa. In co-cultures, M. aeruginosa won in red light and white light, while C. pyrenoidosa dominated under blue light. Light color mainly affected the absorption flux of reaction center (ABS/RC) in photosynthetic system II (PSII) and its potential photosynthetic capacity (Fv/Fm). Fv/Fm of M. aeruginosa in red light (or C. pyrenoidosa in blue light) was significantly enhanced. This study revealed that light color showed a significant influence on interspecific competition between green algae and cyanobacteria, which offers new insights into the dominance establishment and bloom formation of Microcystis.

Extracellular polysaccharide (EPS) can be distinguished into soluble or bound types and significantly contributes to colony formation in Microcystis. Depending on the binding strength with cells, the bilayer structure of bound EPS contains loosely or tightly bound EPS (LB-EPS or TB-EPS) and their roles in shaping the size and tightness of Microcystis colonies deserve further investigation. In this study, the influences of two types of bound EPS on the size and tightness of Microcystis colonies were investigated after a series of pretreatment to obtain LB-EPS retaining or stripped samples. Results showed that cells with LB-EPS formed large and loose colonies. Furthermore, the ratios of LB-EPS to TB-EPS, which indicate the size and tightness of the colonies, were higher in the retaining groups than in the stripped groups. Our findings also provide evidence that calcium enrichment is conducive to colony formation in Microcystis. This study provides new insights into the formation and enlargement of Microcystis colonies, which contributes to a better understanding on the role of EPS in Microcystis aggregation and morphology changes.

Many bat pups emit isolation calls with low-frequency components that facilitate maternal recognition. Most previous studies have noted that isolation calls generally have unique individual identit...

Bats are altricial, gregarious, and highly vocal animals. As such, they are compelling animal models for bioacoustic analyses of language-related traits. Insight into the vocal development of bats ...

Zika virus (ZIKV) infection during gestation is deemed to be coupled to birth defects through direct impairment of the nervous system during neurogenesis. However, in this study, our data showed that ZIKV infection dramatically suppressed cranial osteogenesis, shown by Safranin O/Fast Green and alizarin red staining, in chick embryos, which provides another possibility that craniofacial bone malformation caused by ZIKV may be a major cause of ZIKV-mediated birth defects. By immunofluorescent staining and electron microcopy, we confirmed ZIKV infection in chick embryo neural tubes and sites of neural crest. Next, in vivo (chick embryos) and in vitro [primary culture of neural crest cells (NCC)] ZIKV and HNK-1 double immunofluorescent staining demonstrated that ZIKV infection inhibited the production of migratory NCC. The reduction of both AP-2α- and Pax7-positive NCC in HH10 chick embryos infected by ZIKV confirmed that abnormal development of cranial NCC also occurred in the migratory process. Whole mount in situ hybridization demonstrated that cadherin 6B expression was elevated and Slug, FoxD3, and BMP4/Msx1 expressions decreased in ZIKV-infected HH10 chick embryos, implying that epithelial-mesenchymal transition (EMT) of neural crest production was blocked by ZIKV infection. Moreover, in vivo and in vitro pHIS3 and Pax7 double immunofluorescent staining showed that NCC proliferation was repressed by ZIKV infection. C-caspase-3 and AP-2α double immunofluorescent staining in HH10 chick embryos and western blotting showed that NCC apoptosis increased following ZIKV infection. Finally, electron microscopy showed multiple autophagosomes in ZIKV-infected embryos, and western blot and LC3B immunofluorescent staining demonstrated that autophagy-related genes were activated by ZIKV infection. Taken together, our data first showed that ZIKV infection during embryogenesis could interfere with cranial neural crest development, which in turn causes aberrant cranial osteogenesis. Our results provided new insights into brain malformations induced by ZIKV infection.

Uveitis is an intraocular inflammatory disease which can lead to serious visual impairment. Genetic factors have been shown to be involved in its development. However, few databases have focused on the information of associations between single nucleotide polymorphisms (SNPs) and uveitis. To discover the exact genetic background of uveitis, we developed an SNP database specific for uveitis, “UVEOGENE,” which includes 370 genes and 918 SNPs covering 14 uveitis entities and 40 populations from 286 PubMed English‐language papers. Stratification analyses by gender, HLA status, and different clinical features were also extracted from the publications. As a result, 371 associations were judged as “statistically significant.” These associations were also shared with Global Variome shared Leiden Open Variation Database (LOVD) (https://databases.lovd.nl/shared/genes). Based on these associations, we investigated the genetic relationship among three widely studied uveitis entities including Behcet's disease (BD), Vogt–Koyanagi–Harada (VKH) disease, and acute anterior uveitis (AAU). Furthermore, “UVEOGENE” can be used as a reliable and informative resource to identify similarities as well as differences in the genetic susceptibility among uveitis and other autoimmune diseases. UVEOGENE is freely accessible at http://www.uvogene.com.

Pico-cyanobacteria and micro-cyanobacteria coexist ubiquitously in many lakes. Differences in cell size and abilities to utilize nutrients may influence their distribution patterns. In this study, Synechococcus sp. and Microcystis aeruginosa were chosen as pico- and micro-cyanobacteria, respectively. Gradient phosphorus treatments (0.002, 0.01, 0.05, and 0.25 mg P L−1) were designed in mono- and co-cultures. Growth curves were recorded and fitted by the Monod equation. Moreover, the interspecific competition was analyzed by the Lotka–Volterra model. When mono-cultured in lower P conditions (≤ 0.01 mg P L−1), Synechococcus sp. obtained much higher biomass than M. aeruginosa. But, M. aeruginosa grew faster than Synechococcus sp. in higher P groups (≥ 0.05 mg P L−1) (p

Recombinant interferon-α (rIFN-α) has been widely used for treating viral infections. However, the clinical efficacy of unmodified rIFN-α is limited due to small molecular size and rapid clearance from circulation. In this study we developed a novel strategy for half-life extension of porcine IFN-α (PoIFN-α) by fusion to the immunoglobulin (Ig)-binding C2 domain of streptococcal protein G (SPG). The coding sequences for PoIFN-α6 and SPG C2 domain, with a tobacco etch virus (TEV) protease recognition sequence introduced at the 5-end, were cloned into an elastin-like polypeptide (ELP) fusion expression vector and expressed as an ELP-PoIFNα-C2 fusion protein. After optimization of the conditions for soluble protein expression and purification, the fusion protein was purified to more than 90% purity by two rounds of inverse transition cycling (ITC) in the presence of 0.5% Triton X-100. After cleavage with self-aggregating peptide ELK-16-tagged tobacco etch virus protease, the protease was removed by quick centrifugation and PoIFNα-C2 protein was recovered by an additional round of ITC with 98% purity. Western blotting analysis showed that PoIFNα-C2 protein had the specific affinity for pig IgG binding. The antiviral assay showed that PoIFNα-C2 protein had potent antiviral activities against vesicular stomatitis virus and porcine pseudorabies virus. After single intravenous or subcutaneous injection into rats, PoIFNα-C2 protein showed 16- or 4-fold increase in serum half-life with significantly improved bioavailability.

The objectives of the present study were to identify key genes and biological pathways associated with thermal stress in Chinese Holstein dairy cattle. Hence, we constructed a cell-model, applied various molecular biology experimental techniques and bioinformatics analysis. A total of 55 candidate genes were screened from published literature and the IPA database to examine its regulation under cold (25°C) or heat (42°C) stress in PBMCs. We identified 29 (3 up-regulated and 26 down-regulated) and 41 (15 up-regulated and 26 down-regulated) significantly differentially expressed genes (DEGs) (fold change ≥ 1.2-fold and P < 0.05) after cold and heat stress treatments, respectively. Furthermore, bioinformatics analyses confirmed that major biological processes and pathways associated with thermal stress include protein folding and refolding, protein phosphorylation, transcription factor binding, immune effector process, negative regulation of cell proliferation, autophagy, apoptosis, protein processing in endoplasmic reticulum, estrogen signaling pathway, pathways related to cancer, PI3K- Akt signaling pathway, and MAPK signaling pathway. Based on validation at the cellular and individual levels, the mRNA expression of the HIF1A gene showed upregulation during cold stress and the EIF2A, HSPA1A, HSP90AA1, and HSF1 genes showed downregulation after heat exposure. The RT-qPCR and western blot results revealed that the HIF1A after cold stress and the EIF2A, HSPA1A, HSP90AA1, and HSF1 after heat stress had consistent trend changes at the cellular transcription and translation levels, suggesting as key genes associated with thermal stress response in Holstein dairy cattle. The cellular model established in this study with PBMCs provides a suitable platform to improve our understanding of thermal stress in dairy cattle. Moreover, this study provides an opportunity to develop simultaneously both high-yielding and thermotolerant Chinese Holstein cattle through marker-assisted selection.

BACKGROUND Nitrogen and water are two major factors in rice production. Due to the lack of ample evidence and much uncertainty in field experiments, the coupling effects of water and nitrogen in paddy fields have remained debatable over recent years. RESULTS A fine-calibrated ORYZA (v3) model was applied to simulate rice growth and development under different nitrogen (N) rates and irrigation regimes for a double rice-cropping system in South China. We designed a numerical experiment of 504 treatments, consisting of seven nitrogen rates (0-300 kg ha-1 ), eight irrigation thresholds (30-100%, presented as the percentage of saturated soil water content) and nine irrigation quotas (20-100 mm), and each treatment was simulated for 30 years. Yield varied greatly with different water-nitrogen conditions, particularly in the scenario of frequently alternate wetting and drying irrigation and low-N rates. The coupling effects had a negligible influence on water input and water loss, which were found to be sensitive only to the irrigation regime and rainfall distribution. Based on the results, the N fertilizer for early rice growing in the wet season is suggested as 150-200 kg ha-1 , and 200-250 kg ha-1 for late rice growing in the dry season. The irrigating threshold and irrigation quota for early rice are suggested as lower than 70% and 30-40 mm, respectively, and, for late rice, 70-80% and 40-60 mm. CONCLUSION Remarkable water-nitrogen coupling effects were found in the paddy field, and integrative water-nitrogen management strategies were suggested for both early rice and late rice in South China. © 2021 Society of Chemical Industry.

Abstract: Kallikrein 7 (KLK7) is a secreted serine protease with chymotrypsic protease activity. Abnormally high expression of KLK7 is closely related to the occurrence and development of various types of cancer. Therefore, KLK7 has been identified as a potential target for cancer drug development design in recent years. KLK7 mediates various biological and pathological processes in tumorigenesis, including cell proliferation, migration, invasion, angiogenesis, and cell metabolism, by hydrolyzing a series of substrates such as membrane proteins, extracellular matrix proteins, and cytokines. This review mainly introduces the downstream cell signaling pathways involved in the activation of KLK7 and its substrate-related proteins. This review will not only help us to better understand the mechanisms of KLK7 in regulating biological and pathological processes of cancer cells but also lay a solid foundation for the design of inhibitors targeting KLK7.

This study analyzed the difference between biofilm and planktonic Brucella abortus using metabolomics and proteomics. Brucella abortus was cultured in different media to induce Brucella abortus biofilm formation and planktonic cells, followed by metabolomics and proteomics analyses for these two samples. Significant differential metabolites were identified, followed by KEGG pathway analysis. Differentially expressed proteins were identified, followed by subcellular localization, GO annotation, and KEGG pathway enrichment. Additionally, a correlation analysis of metabolomics and proteomics was performed. Metabolomics analysis showed 7682 positive and 4433 negative metabolites, including 188 positive and 117 negative significant differential metabolites. These differential metabolites were enriched in fatty acid/unsaturated fatty acid biosynthesis and linoleic acid metabolism. Proteomics analysis revealed 1759 proteins, including 486 differentially expressed proteins, which were enriched in various metabolic and degradation-related pathways. Subcellular localization showed that 74.3% of the differential proteins were cytoplasmic proteins. Correlation analysis showed that 1-palmitoyl-2-oleoyl-phosphatidylglycerol had the most significant correlations with proteins, followed by cytosine. Both metabolites correlated with the protein Q57EI7 (RbsB-1, ribose ABC transporter). One common pathway, fatty acid biosynthesis, was identified by both proteomics and metabolomics analyses that involved the metabolites, oleic acid, and protein Q57DK3 (biotin carboxylase). There were metabolomic and proteomic differences between Brucella abortus biofilm and planktonic cells, and these results provide novel insights into the biofilm-forming process of Brucella abortus.

Trait-based approaches provide a mechanistic framework crossing scales from cellular traits to community dynamics, while ecological stoichiometry applies first principles to understand how the balance of energy and elements shape ecological interactions. However, few studies have explicitly linked both frameworks. In this study, we tested the stoichiometric regulation of a number of carbon (C) based (e.g., extracellular polysaccharides and colony formation) and nitrogen (N) containing traits (i.e., chlorophyll a, phycocyanin, and gas vesicle content) in cyanobacteria in laboratory experiments and in the field. We exposed the cosmopolitan colony forming freshwater cyanobacterium Microcystis sp. in batch experiments to light, N and phosphorus (P) limitation, and enhanced CO2 levels, and assessed the regulation of these traits. Cyanobacterial traits followed stoichiometrically predictable patterns, where N containing traits increased with cellular N content, and decreased with increasing C : N ratios. C-based traits increased with cellular C content and C : N ratios under nutrient, particularly N, limitation. The pattern of colony formation was confirmed with field data from Lake Taihu (China), showing an increase in colony size when N was limiting and N : P ratios were low. These findings demonstrate how an explicit coupling of trait-based approaches to ecological stoichiometry can support our mechanistic understanding of responses of cyanobacteria toward shifts in resource availability.

The clinical significance of the family with sequence similarity 189 member B (FAM189B) gene remains largely unknown in gastric cancer (GC). A comprehensive investigation combining multiple detection methods was carried out in the current study to unveil the clinical implications and prospective molecular characterization of FAM189B protein and mRNA in GC. The protein level of FAM189B was clearly upregulated in the tumor tissues of GC as compared to noncancerous gastric tissues with 179 GC cases and 147 noncancerous gastric controls assessed by immunohistochemistry. The upregulation of the FAM189B protein was also found in the more deteriorating period of the tumor, as there were increasing trends in the groups of larger tumors, with lymph node metastasis, a further advanced clinical stage, and a higher histological grade. Next, we focused on the mRNA level of FAM189B in GC tissues using various high-throughput data. After the screening of GEO, ArrayExpress, and SRA, we finally achieved 18 datasets, including an RNA sequencing dataset of TCGA. Altogether, 1095 cases of GC tissue samples were collected, with 305 unique examples of noncancerous controls. Concerning the mRNA level of FAM189B in GC, the final standard mean difference (SMD) was 0.46 and the area under the curve (AUC) was 0.79 for the upregulation of FAM189B mRNA, which confirmed that the FAM189B mRNA level was also markedly upregulated in GC tissues and comparable to its protein level. The survival analysis showed that the higher expression of FAM189B was a risk factor for the overall survival, first progression, and postprogression survival of GC. For the Affymetrix ID 1555515_a_at of FAM189B, the higher expression level of FAM189B predicted a lower overall survival, first progression survival, and postprogression survival with the hazard ratio (HR) being 1.56 (1.24, 1.95), 1.69 (1.32, 2.17), and 1.97 (1.5, 2.6), respectively. For the Affymetrix ID 203550_s_at of FAM189B, a similar result could be found with corresponding HR being 1.49 (1.24, 1.8), 1.49 (1.21, 1.83), and 1.66 (1.32, 2.08), respectively. The interaction of MEM, COXPRESdb coexpressed genes, and DEGs of GC finally generated 368 genes, and the pathway of the cell cycle was the top pathway enriched by KEGG. In conclusion, the overexpression of the FAM189B protein and mRNA might enhance the incidence of GC.

Although the survival rate of patients with cancer have increased due to the use of current chemotherapeutic agents, adverse effects of cancer therapy remain a concern. The reversal of drug resistance, reduction in harmful side effects and accelerated increase in efficiency have often been addressed in the development of combination therapeutics. Tazemetostat (EPZ-6438), a histone methyltransferase EZH2 selective inhibitor, was approved by the FDA for the treatment of advanced epithelioid sarcoma. However, the effect of tazemetostat on colorectal cancer (CRC) and 5-FU sensitivity remains unclear. In this study, the enhancement of tazemetostat on 5-FU sensitivity was examined in CRC cells. Our findings demonstrated that tazemetostat combined with 5-FU exhibits synergistic antitumor function in vitro and in vivo in CRC cells. In addition, tazemetostat promotes PUMA induction through the ROS/ER stress/CHOP axis. PUMA depletion attenuates the antitumor effect of the combination therapy. Therefore, tazemetostat may be a novel treatment to improve the sensitivity of tumors to 5-FU in CRC therapy. In conclusion, the combination of 5-FU and tazemetostat shows high therapeutic possibility with reduced unfavorable effects.

Understanding the molecular mechanisms underlying mammalian kidney function requires transcriptome profiling of the interplay between cells comprising nephron segments. Traditional transcriptomics requires cell dissociation, resulting in loss of the spatial context of gene expression within native tissue. To address this problem, we performed spatial transcriptomics (ST) to retain the spatial context of the transcriptome in human and mouse kidneys. The generated ST data allowed spatially resolved differential gene expression analysis, spatial identification of functional nephron segments, cell-to-cell interaction analysis, and chronic kidney disease-associated genetic variant calling. Novel ST thus provides an opportunity to enhance kidney diagnostics and knowledge, by retaining the spatial context of gene expression within intact tissue.

PurposeThe liver is the most frequent metastatic site for colorectal cancer (CRC). Its microenvironment is modified to provide a niche that allows CRC cell growth. This study focused on characterizing the cellular changes in the metastatic CRC (mCRC) liver tumor microenvironment (TME).Experimental DesignWe analyzed a series of microsatellite stable (MSS) mCRCs to the liver, paired normal liver tissue and peripheral blood mononuclear cells using single cell RNA-seq (scRNA-seq). We validated our findings using multiplexed spatial imaging and bulk gene expression with cell deconvolution.ResultsWe identified TME-specific SPP1-expressing macrophages with altered metabolism features, foam cell characteristics and increased activity for extracellular matrix (ECM) organization. SPP1+ macrophages and fibroblasts expressed complementary ligand receptor pairs with the potential to mutually influence their gene expression programs. TME lacked dysfunctional CD8 T cells and contained regulatory T cells, indicative of immunosuppression. Spatial imaging validated these cell states in the TME. Moreover, TME macrophages and fibroblasts had close spatial proximity, a requirement for intercellular communication and networking. In an independent cohort of mCRCs in the liver, we confirmed the presence of SPP1+ macrophages and fibroblasts using gene expression data. An increased proportion of TME fibroblasts was associated with worst prognosis in these patients.ConclusionsWe demonstrated that mCRC in the liver is characterized by transcriptional alterations of macrophages in the TME. Intercellular networking between macrophages and fibroblasts supports CRC growth in the immunosuppressed metastatic niche in the liver. These features can be used to target these immune checkpoint resistant MSS tumors.TRANSLATIONAL RELEVANCEThe liver is the commonest site for metastatic colorectal cancer (mCRC). Alterations in the tumor microenvironment (TME) allow metastatic cells to seed the distant liver site and grow. Leveraging single-cell RNA sequencing, we discovered a distinct SPP1+ macrophage cell state with pro-fibrogenic gene expression and altered metabolism. These SPP1+ macrophages communicated with fibroblasts, mutually influencing each other’s gene expression program. Using spatial imaging, we confirmed proximal colocalization between macrophages and fibroblasts in the mCRC TME, which is required for intercellular communication. These states and intercellular communication promoted immunosuppression in the TME, with a lack of dysfunctional anti-tumor CD8 T cells and prevalence of regulatory T cells. Increased fibroblasts were associated with worst prognosis in an independent patient cohort. Our results identified novel TME features that result in reshaping of the metastatic niche that allows progression of mCRC. These features can be potential targets for mCRC treatment, which is microsatellite stable and resistant to immune checkpoint blockade.

Single nucleotide polymorphisms (SNPs) in the IL1RL1-IL18R1 region are associated with various immune-mediated diseases. This study was carried out to investigate the causal variant for ocular Behcet's disease (BD) and elucidate its target genes in the IL1RL1-IL18R1 region. Nine candidate functional SNPs were prioritized with bioinformatics analysis, followed by a two-stage association study in 694 ocular BD patients and 1,458 unaffected controls. Functional studies were performed in the peripheral blood mononuclear cells (PBMCs) of 45 healthy men and 16 active male BD patients. Genotyping was performed using the MassARRAY System. The mRNA expressions of IL1RL1, IL18R1, IL18RAP, and SLC9A4 were assayed by real-time PCR and secretion of cytokines was examined by ELISA. Significantly lower frequencies of the rs12987977 GG genotype/G allele (P-c= 8.93 x 10(-7), OR = 0.39;P-c= 2.60 x 10(-3), OR = 0.77, respectively), rs12999364 TT genotype/T allele (P-c= 3.15 x 10(-4), OR = 0.51;P-c= 1.13 x 10(-2), OR = 0.80, respectively), and rs4851569 AA genotype/A allele (P-c= 3.29 x 10(-4), OR = 0.52;P-c= 9.72 x 10(-3), OR = 0.80, respectively) were observed in BD patients compared with the controls. Functional experiments revealed a downregulation of IL1RL1, IL18R1, and SLC9A4 and a decreased secretion of IFN-gamma in the anti-CD3/CD28 antibody-treated PBMCs as well as a decreased production of TNF-alpha in the lipopolysaccharide (LPS)-stimulated PBMCs in carriers of the protective homozygous rs12987977/GG genotype compared with the TT genotype. Our findings show that functional SNPs-rs12987977, rs12999364, and rs4851569-in the IL1RL1-IL18R1 region confer susceptibility to ocular BD in a Chinese Han population. And IL1RL1, IL18R1, and SLC9A4 may be the target genes of rs12987977.

Spatial Transcriptomics is an emerging technology that adds spatial dimensionality and tissue morphology to the genome-wide transcriptional profile of cells in an undissociated tissue. Integrating these three types of data creates a vast potential for deciphering novel biology of cell types in their native morphological context. Here we developed innovative integrative analysis approaches to utilise all three data types to first find cell types, then reconstruct cell type evolution within a tissue, and search for tissue regions with high cell-to-cell interactions. First, for normalisation of gene expression, we compute a distance measure using morphological similarity and neighbourhood smoothing. The normalised data is then used to find clusters that represent transcriptional profiles of specific cell types and cellular phenotypes. Clusters are further sub-clustered if cells are spatially separated. Analysing anatomical regions in three mouse brain sections and 12 human brain datasets, we found the spatial clustering method more accurate and sensitive than other methods. Second, we introduce a method to calculate transcriptional states by pseudo-space-time (PST) distance. PST distance is a function of physical distance (spatial distance) and gene expression distance (pseudotime distance) to estimate the pairwise similarity between transcriptional profiles among cells within a tissue. We reconstruct spatial transition gradients within and between cell types that are connected locally within a cluster, or globally between clusters, by a directed minimum spanning tree optimisation approach for PST distance. The PST algorithm could model spatial transition from non-invasive to invasive cells within a breast cancer dataset. Third, we utilise spatial information and gene expression profiles to identify locations in the tissue where there is both high ligand-receptor interaction activity and diverse cell type co-localisation. These tissue locations are predicted to be hotspots where cell-cell interactions are more likely to occur. We detected tissue regions and ligand-receptor pairs significantly enriched compared to background distribution across a breast cancer tissue. Together, these three algorithms, implemented in a comprehensive Python software stLearn, allow for the elucidation of biological processes within healthy and diseased tissues.

Age-related macular degeneration and Alzheimer's disease are closely related complex diseases that may share overlapping pathogenesis in gene networks. This study was conducted to investigate the genetic factors shared by both diseases. We analyzed genome-wide association studies' summary statistics from both diseases using a new platform known as functional mapping and annotation (FUMA) and a co-localization analysis. We obtained disease-related gene expression profile data from the Gene Expression Omnibus and analyzed these data using weighted gene co-expression network analysis. FUMA analysis and Bayesian co-localization analysis showed that ten genes on chromosome 7, one pathway (complement and coagulation cascade), and forty-two biological processes were common for both diseases. Among these ten genes, two protein-coding genes, two pseudogenes, and two RNA genes were, for the first time, identified to be associated with both diseases. Weighted gene co-expression network analysis identified 19 age-related macular degeneration hub genes and 19 Alzheimer's disease hub genes and revealed that these two diseases shared nine pathways and 63 biological processes. Using FUMA, co-localization analysis, and weighted gene co-expression network analysis, 10 genes on chromosome 7, 10 pathways, and 105 biological processes were found to be associated with the two degenerative diseases. This suggests that these10 genes and the hub genes of these modules associated with the shared pathways are potential diagnostic markers for both diseases.

In water treatment process, Microcystis colonies can be effectively removed by coagulants. However, the use of popular coagulants could cause adverse health effects in humans or increase the amount of sludge. Meanwhile, Microcystis unicells are much more difficult to remove than colonies, due to their small size and dispersed distribution. This study proposed and analyzed the flocculation of Microcystis unicells induced by pH regulation. The particle size, zeta potential, cell viability and integrity, cytochemical changes, and cell-to-cell connections were recorded during pH regulation. Results showed that when pH was adjusted in the range of 2.5 to 2 by HCl (1.2 M), Microcystis unicells aggregated to form flocs as large as 28 μm, which are easy to remove by filtration or sedimentation. The overwhelming majority of cells were intact and inactivated in the optimal pH range (2.5–2). Thus, pH regulation is an environment-friendly and cost-effective method to remove Microcystis unicells, which can be potentially applied to water treatment.

Baicalin is a flavone glycoside that possesses numerous pharmacological properties. but its protective mode of action in kidney injury induced by diabetes mellitus remains incompletely understood. Using a streptozotocin (STZ)-induced diabetic mouse model, we found that baicalin could ameliorate diabetes-induced the pathological changes of the kidney function and morphology through suppressing inflammation and oxidative stress. Furthermore, baicalin treatment could alleviate interstitial fibrosis in the diabetic kidney via inhibiting epithelial-to-mesenchymal transition (EMT), which was accompanied by a sharp upregulation of Klotho, the endogenous inhibitor of renal fibrosis. We further verified that baicalin-rescued expression of Klotho was associated with Klotho promoter hypomethylation due to aberrant methyltransferase 3a expressions. Klotho knockdown via RNA interferences largely abrogated the anti-renal fibrotic effects of Baicalin in HK2 cells. These findings suggested that baicalin could alleviate renal injury-induced by diabates through partly modulating Klotho promoter methylation, which provides new insights into the treatment of diabetic nephropathy.

Mycobacterium tuberculosis (Mtb), a typical intracellular parasite that cause tuberculosis (TB), which rank the top infectious killer for human death. Pathogenic mycobacterium has evolved numerous strategies to favor their intracellular survival including a unique lipid-rich cell wall component and granuloma formation during early infection [1]. Phthiocerol dimycocerosate (PDIM) is a critical virulence factor of mycobacteria including Mycobacterium marinum (Mm), which manipulate host immune responses and granuloma induction and facilitate immune evasion [2-7], but the mechanisms are still remains questionable. We used an AACT/SILAC-based quantitative proteomic approach to determine PDIM-responded proteome profiles during Mm early infection. A major difference was the high abundance of galectin3 (Gal-3) in WT Mm-infected cells and not with PDIM-deficient. Gal-3 induction by PDIM-replete bacteria was primarily via Toll-like receptor 2, and also engaged TGF-β non-classical pathway. Elevated Gal-3 in macrophages prevented the turnover and translocation of NF-κB to effectively modulate the profile of inflammatory cytokines. Silencing of Gal-3 effectively relieved tissue damage induced in mice by PDIM-expressing Mm. We found upregulated Gal-3 in the serum and BALF of confirmed clinical tuberculosis cases, while decreased significantly after effective chemotherapy. Our findings demonstrated that upregulated Gal-3 not only plays an important role in regulating host immune response and granuloma formation in vivo, but also that target-Gal-3 therapy is a promising anti-tuberculosis strategy in the future. Funding Statement: This work was supported in part by the National Natural Science Foundation of China (Grant Nos. 31772709 and 31572485 to D.W), the new faculty startup research fund of China Three Gorges University (KJ2014B023 to D.W), Health Commission of Hubei Province in China (WJ2019H528 to D.W; WJ2019F080 to D.L), and in part by the intramural research program of NIAID, NIH (Bethesda MD, USA). Z.L was supported by grants from the China’s 13th Five Year Programs for the prevention and cure of great infectious diseases (2017ZX10201301-005) and Open Fund of Shanghai Key Laboratory of Tuberculosis (2018kf03). XY.Fan is the recipient of National Natural Science Foundation of China (31771004). Declaration of Interests: The authors declare no competing financial interests. Ethics Approval Statement: The case-control study of TB was reviewed and approved by the Ethics Committee of the Third People’s Hospital of Yichang (2017008H). Written informed consent was obtained from all participants. The animal experiments were reviewed and approved by the Animal Care and Use Committee of China Three Gorges University (2017010M).

Well-preserved Ginkgo leaves with cuticle were collected from the Middle Jurassic Jiulongshan Formation in the Daohugou area, Inner Mongolia, China, which form the common elements in the Daohugou flora. Three new species of Ginkgo are recognized and their leaf morphology and cuticular structure are described for the first time. Two different pCO2 proxy models were applied to reconstruct palaeo-atmospheric CO2 concentration by using these new Ginkgo material. NLE (nearest living equivalent) suggests a semi-quantitative pCO2 estimate as 839 ± 99 ppmv with Carboniferous standardization and 419 ± 49 ppmv with Recent standardization. While Barclay’s revised SI-pCO2 regression suggests the estimates as 405 ± 71 ppmv. The results show that the pCO2 estimate is higher than today’s atmospheric CO2 concentration but lower than most of the other results on the Jurassic Period. Combined with the floristic composition, the climate at 165 Ma in the Daohugou area is inferred to be warm temperate.

SummaryFibroblast growth factor (FGF) signalling acts as one of modulators that control neural crest cell (NCC) migration, but how this is achieved is still unclear. In this study, we investigated the effects of FGF signalling on NCC migration by blocking this process. Constructs that were capable of inducing Sprouty2 (Spry2) or dominant-negative FGFR1 (Dn-FGFR1) expression were transfected into the cells making up the neural tubes. Our results revealed that blocking FGF signalling at stage HH10 (neurulation stage) could enhance NCC migration at both the cranial and trunk levels in the developing embryos. It was established that FGF-mediated NCC migration was not due to altering the expression of N-cadherin in the neural tube. Instead, we determined that cyclin D1 was overexpressed in the cranial and trunk levels when Sprouty2 was upregulated in the dorsal neural tube. These results imply that the cell cycle was a target of FGF signalling through which it regulates NCC migration at the neurulation stage.

To explore the possible roles of metallothioneins (MTs) played in cadmium (Cd) accumulation of water spinach, three IaMT genes, IaMT1, IaMT2 and IaMT3 in a high-shoot-Cd (T308) and a low-shoot-Cd accumulation cultivar (QLQ) were cloned, characterized, and quantitated. Gene expression analysis suggested that the expression of the IaMTs was differentially regulated by Cd stress in different cultivars, and T308 showed higher MTs expression overall. Furthermore, only shoot IaMT3 expression was cultivar dependent among the three IaMTs. Antioxidant analysis showed that the high production of IaMTs in T308 should be associated with its high oxidation resistance. The role of IaMTs in protecting against Cd toxicity was demonstrated in vitro via recombinant E. coli strains. The results showed that IaMT1 correlated with neither Cd tolerance nor Cd accumulation of E. coli, while IaMT2 conferred Cd tolerance in E. coli, IaMT2 and IaMT3 increased Cd accumulation in E. coli. These findings help to clarify the roles of IaMTs in Cd accumulation, and increase our understanding of the cultivar-dependent Cd accumulation in water spinach.

This study aims at investigating the radioprotective effect of ethanol extract from Ji-Xue-Teng (JXT,Spatholobus suberectus) on radiation-induced hematopoietic alteration and oxidative stress in the liver. Mice were exposed to a single acuteγ-radiation for the whole body at the dose of 6.0 Gy, then subjected to administration of amifostine (45 mg/kg) or JXT (40 g crude drug/kg) once a day for 28 consecutive days, respectively. Bone marrow cells and hemogram including white cells, red cells, platelet counts, and hemoglobin level were examined. The protein expression levels of pJAK2/JAK2, pSTAT5a/STAT5a, pSTAT5b/STAT5b, and Bcl-2 in bone marrow tissue; levels of reactive oxygen species (ROS); and the activity of superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione peroxidase (GSH-Px) in serum and liver tissue were determined. At the end of the experiment, the effect of JXT on cell viability and G-CSF and G-CSFR levels in NFS-60 cells were tested by CCK-8 assay, ELISA, and flow cytometry. The results showed that the mice exposed toγ-radiation alone exhibited a typical hematopoietic syndrome. In contrast, at the end of the 28-day experiment, irradiated mice subjected to oral administration of JXT showed an obvious improvement on blood profile with reduced leucopenia, thrombocytopenia (platelet counts), RBC, and hemoglobin levels, as well as bone marrow cells. The expression of pJAK2/JAK2, pSTAT5a/STAT5a, and Bcl-2 in bone marrow tissue was increased after JXT treatment. The elevation of ROS was due to radiation-induced toxicity, but JXT significantly reduced the ROS level in serum and liver tissue, elevated endogenous SOD and GSH-PX levels, and reduced the MDA level in the liver. JXT could also increase cell viability and G-CSFR level in NFS-60 cells, which was similar to exogenous G-CSF. Our findings suggested that oral administration of JXT effectively facilitated the recovery of hematopoietic bone marrow damage and oxidative stress of the mice induced byγ-radiation.

GNA13 has been found overexpressed in various types of cancer, which is related to tumor metastasis and progression. However, the biological functions of GNA13 in colorectal cancer (CRC) progression remain unclear. This study aimed to explore the role of GNA13 in CRC and investigate the mechanism of how GNA13 promotes tumor growth. Interestingly, our findings showed that GNA13 is commonly upregulated in CRC, where these events are associated with a worse histologic grade and poor survival. Increased expression levels of GNA13 promoted cell growth, migration, invasion, and epithelial‐mesenchymal transition, whereas GNA13 silencing abrogated these malignant phenotypes. In addition, overexpressing GNA13 in cancer cells increased the levels of the chemokines CXCL1, CXCL2, and CXCL4, which contributed to CRC proliferation and colony formation. Moreover, our mechanistic investigations suggest that the NF‐κB/p65 signaling pathway was activated by the increase in GNA13 levels. Inhibiting the NF‐κB/p65 pathway with an inhibitor decreased GNA13‐induced migration, invasion and CXCL chemokine level increases, indicating the critical role of NF‐κB/p65 signaling in mediating the effects of GNA13 in CRC. Together, these results demonstrate a key role of GNA13 overexpression in CRC that contributes to malignant behavior in cancer cells, at least in part through stimulating angiogenesis and increasing the levels of the NF‐κB‐dependent chemokines CXCL1, CXCL2, and CXCL4.

The clinical use of recombinant interferons (rIFNs) is limited by higher purification cost and quick clearance from circulation. Elastin-like polypeptides (ELPs) are a novel tag for recombinant protein purification and half-life extension. In this study, we evaluated the feasibility of ELP fusion for simple purification and half-life extension of recombinant porcine IFNs (rPoIFNs). After construction of five different fusion expression vectors, we optimized the conditions for soluble protein expression and purification. SDS-PAGE analysis showed that, unlike PoIFNα-His and PoIFNγ-His, PoIFNα-ELP, ELP-PoIFNα and PoIFNαγ-ELP were expressed mainly as soluble proteins at 20 ℃. The optimal conditions for the inverse transition cycling (ITC) of three ELP fusion proteins were 2 M NaCl at 28 ℃. After two rounds of ITC, the three ELP fusion proteins were purified to more than 90% purities, which were comparable to that of affinity-purified PoIFNα-His and PoIFNγ-His. Cytopathic effect inhibition assay showed that the five rPoIFNs had potent but different antiviral activities against two different viruses on two different cell types. The plasma solubility assay showed that the three ELP-fused rPoIFNs remained as soluble proteins under the physical conditions. The plasma stability of three ELP-fused rPoIFNs was significantly improved in comparison with that of PoIFN-α. These data suggest that ELP fusion is a feasible strategy to enhance purification and plasma stability of rPoIFNs.

Both Xp11 translocation renal cell carcinomas and the corresponding mesenchymal neoplasms are characterized by a variety of gene fusions involving TFE3. It has been known that tumors with different gene fusions may have different clinicopathologic features; however, further in-depth investigations of subtyping Xp11 translocation-associated cancers are needed in order to explore more meaningful clinicopathologic correlations. A total of 22 unusual cases of Xp11 translocation-associated cancers were selected for the current study; 20 cases were further analyzed by RNA sequencing to explore their TFE3 gene fusion partners. RNA sequencing identified 17 of 20 cases (85%) with TFE3-associated gene fusions, including 4 ASPSCR1/ASPL-TFE3, 3 PRCC-TFE3, 3 SFPQ/PSF-TFE3, 1 NONO-TFE3, 4 MED15-TFE3, 1 MATR3-TFE3, and 1 FUBP1-TFE3. The results have been verified by fusion fluorescence in situ hybridization (FISH) assays or reverse transcriptase polymerase chain reaction (RT-PCR). The remaining 2 cases with specific pathologic features highly suggestive of MED15-TFE3 renal cell carcinoma were identified by fusion FISH assay. We provide the detailed morphologic and immunophenotypic description of the MED15-TFE3 renal cell carcinomas, which frequently demonstrate extensively cystic architecture, similar to multilocular cystic renal neoplasm of low malignant potential, and expressed cathepsin K and melanotic biomarker Melan A. This is the first time to correlate the MED15-TFE3 renal cell carcinoma with specific clinicopathologic features. We also report the first case of the corresponding mesenchymal neoplasm with MED15-TFE3 gene fusion. Additional novel TFE3 gene fusion partners, MATR3 and FUBP1, were identified. Cases with ASPSCR1-TFE3, SFPQ-TFE3, PRCC-TFE3, and NONO-TFE3 gene fusion showed a wide variability in morphologic features, including invasive tubulopapillary pattern simulating collecting duct carcinoma, extensive calcification and ossification, and overlapping and high columnar cells with nuclear grooves mimicking tall cell variant of papillary thyroid carcinoma. Furthermore, we respectively evaluated the ability of TFE3 immunohistochemistry, TFE3 FISH, RT-PCR, and RNA sequencing to subclassify Xp11 translocation-associated cancers. In summary, our study expands the list of TFE3 gene fusion partners and the clinicopathologic features of Xp11 translocation-associated cancers, and highlights the importance of subtyping Xp11 translocation-associated cancers combining morphology, immunohistochemistry, and multiple molecular techniques.

Yanliaoa is a common fossil in the Middle Jurassic of western Liaoning, eastern Inner Mongolia and northern Hebei Province, China. It is an important element of the Yanliao biota. The genus was established by Pan in 1977 for fossil plants from the Middle Jurassic Haifanggou Formation in Xiasanjiaochengzi, western Liaoning Province, and in present paper, the genus Yanliaoa is studied based on new material. Pan never designated a type specimen and his fossil material cannot be located. We designate a type specimen here for Yanliaoa, so that the genus name Yanliaoa remains valid. Yanliaoa sinensis Pan emend. Tan et al., is found in the same locality and formation as the lost specimens, Y. sinensis of Pan, 1977. Yanliaoa daohugouensis n. sp., a new species with epidermal anatomy, is from the Middle Jurassic Daohugou, Inner Mongolia. A holotype is also selected from the new material for this new species. Characters of the leafy shoots and ovulate cones of Yanliaoa are emended. The epidermal anatomy of this genus is described for the first time. Compared with other extant and extinct species of Cupressaceae s. l., the current species can be distinguished from any known species both by the leafy shoot characters and its epidermal anatomy. It further indicates that Yanliaoa is an extinct and endemic conifer found in the Middle Jurassic of northeastern China.

Mcl-1, a prosurvival Bcl-2 family protein, is frequently overexpressed in cancer cells and plays a critical role in therapeutic resistance. It is well known that anticancer agents induce phosphorylation of Mcl-1, which promotes its binding to E3 ubiquitin ligases and subsequent proteasomal degradation and apoptosis. However, other functions of Mcl-1 phosphorylation in cancer cell death have not been well characterized. In this study, we show in colon cancer cells that histone deacetylase inhibitors (HDACi) induce GSK3β-dependent Mcl-1 phosphorylation, but not degradation or downregulation. The in vitro and in vivo anticancer effects of HDACi were dependent on Mcl-1 phosphorylation and were blocked by genetic knock-in of a Mcl-1 phosphorylation site mutant. Phosphorylation-dead Mcl-1 maintained cell survival by binding and sequestering BH3-only Bcl-2 family proteins PUMA, Bim, and Noxa, which were upregulated and necessary for apoptosis induction by HDACi. Resistance to HDACi mediated by phosphorylation-dead Mcl-1 was reversed by small-molecule Mcl-1 inhibitors that liberated BH3-only proteins. These results demonstrate a critical role of Mcl-1 phosphorylation in mediating HDACi sensitivity through a novel and degradation-independent mechanism. These results provide new mechanistic insights on how Mcl-1 maintains cancer cell survival and suggest that Mcl-1–targeting agents are broadly useful for overcoming therapeutic resistance in cancer cells. Significance: These findings present a novel degradation–independent function of Mcl-1 phosphorylation in anticancer therapy that could be useful for developing new Mcl-1–targeting agents to overcome therapeutic resistance. Cancer Res; 78(16); 4704–15. ©2018 AACR.

Mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) integrates various environmental signals to regulate cell growth and metabolism. DEPTOR, also termed DEPDC6, is an endogenous inhibitor of mTORC1 and mTORC2 activities. The abundance of DEPTOR centrally orchestrates the mTOR signaling network. However, the mechanisms by which DEPTOR stability is regulated are still elusive. Here, we report that OTU domain–containing ubiquitin aldehyde-binding protein 1 (OTUB1) specifically deubiquitinates DEPTOR in a deubiquitination assay. We found that OTUB1 directly interacted with DEPTOR via its N-terminal domain, deubiquitinated DEPTOR, and thereby stabilized DEPTOR in a Cys-91–independent but Asp-88–dependent manner, suggesting that OTUB1 targets DEPTOR for deubiquitination via a deubiquitinase activity–independent non-canonical mechanism. The interaction between OTUB1 and DEPTOR was enhanced when the cells were treated with amino acids. Moreover, OTUB1 suppressed amino acid–induced activation of mTORC1 in a DEPTOR-dependent manner and thereby ultimately controlled cellular autophagy, cell proliferation, and size. Our findings reveal a mechanism that stabilizes the mTORC1 inhibitor DEPTOR via OTUB1's deubiquitinase activity. Our insights may inform research into various mTOR activity–related diseases, such as cancer, and may contribute to the identification of new diagnostic markers and therapeutic strategies for cancer treatments.

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated nuclease (Cas) based biosensing system provides a novel genomic diagnostic tool for pathogenic detection. However, most of the discovered Cas effectors have poor single strand DNA (ssDNA) target recognition capability with the constraint of protospacer adjacent motif (PAM) sites, which are not suitable for universal pathogenic diagnosis. Herein, we developed a highly sensitive and specific fluorescence tool for bacterial detection by utilizing the unique collateral cleavage activity of a Cas14a1-mediated nucleic acid detection platform (CMP). We combine CMP with molecular amplification to build a CRISPR-Cas based bioanalysis technique, offering fast nucleic acid detection with high sensitivity and specificity. This technique can identify different species of pathogens in milk samples with excellent accuracy. The CMP technique is a promising platform for pathogenic genomic diagnostic in biomedicine and food safety field.

AimTo elucidate the role of microRNA-20a-5p (miR-20a-5p) in the pathogenesis of Vogt-Koyanagi-Harada (VKH) disease.MethodsQuantitative real-time PCR was used to quantify miR-20a-5p expression in CD4+ T cells from patients with active VKH and normal controls. The promoter methylation status of miR-20a-5p was detected by bisulfite sequencing PCR. Targets were evaluated by a luciferase reporter assay. The functional effects of miR-20a-5p on CD4+ T cells from patients with active VKH were assessed by upregulation or downregulation of its expression using liposomes.ResultsThe miR-20a-5p level was significantly decreased in CD4+ T cells from patients with active VKH as compared with normal controls. The two genes, oncostatin M (OSM) and C-C motif chemokine ligand 1 (CCL1), were identified as targets of miR-20a-5p. The upregulation of miR-20a-5p significantly suppressed interleukin 17 (IL-17) production in CD4+ T cells from patients with active VKH, whereas downregulation of miR-20a-5p exhibited an inverse effect. In addition, overexpression of OSM and CCL1 could rescue the effect of the upregulation of miR-20a-5p. Moreover, the level of miR-20a-5p was reduced in response to hypermethylation of the promoter. Further study showed that miR-20a-5p suppressed the activity of the phosphoinositide 3-kinase-AKT pathway.ConclusionsOur findings indicate that downregulation of miR-20a-5p is caused by promoter hypermethylation. MiR-20a-5p could also suppress the production of IL-17 by targeting OSM and CCL1 production in CD4+ T cells in patients with active VKH.

The tree shrew has been used as a primate animal model in neuroscience studies but it has only rarely been employed in the study of reproductive systems. This is mainly because we know very little about the histological features of reproductive organs of the tree shrew. In this study, we have systematically analyzed the histology of reproductive organs of tree shrew, in comparison with human organs. The uterus of female tree shrew is uterus biomes unicolis, which is connected with an enveloped ovary through a thin fallopian tube. Histologically, the fallopian tube consists of folded mucosa, muscularis and serosa. Like other mammalian animals, the different developmental stages (primordial, primary, secondary and Graafian follicles) of ovarian follicles including inner oocyte and outer granulosa cells are embedded in the cortex. The luminal endometrium, middle muscular myometrium and serosa constitute the wall of uterus of tree shrew. The uterine endometrium contains simple columnar ciliated cells and goblet cells, and there are rich uterine glands in underlying stroma. Furthermore, these glands of tree shrew are round and smaller during anestrus, and become much longer when they are in estrus. The uterine endometrium in younger animals was less developed when compared to a mature tree shrew. Compared to human uterine endometrium, the histological features of tree shrew are very similar, indicating that it could potentially be good primate animal model for studying the diseases in reproductive system.

Ultrasonication has drawn an increasing attention as one of cell disruption methods for extracting cellular compounds or controlling algal blooms. However, the effects of biological and physical properties of microalgae on cell disruption were not well understood. In this work, cell disruption of six microalgae, namely, Chlamydomonas reinhardtii, Chlorella pyrenoidosa, Microcystis aeruginosa (three strains: PCC 7806, FACHB 469, and FACHB 1343), and Synechococcus elongatus, was compared mutually based on their characteristics induced by a low-frequency ultrasound (35 kHz, 0.043 W mL−1). Results showed that the most sensitive strain was C. reinhardtii which has a hydroxyproline-rich-glycoproteins cell wall and a larger cell size (normally 10 μm in diameter). More than 80% of the cells of C. reinhardtii were ruptured after sonication for 5 min. In comparison, C. pyrenoidosa, a cellulose-rich-wall algal species with a medium size of 4–6 μm, and M. aeruginosa FACHB 1343, a peptidoglycan-wall species with a smaller average size of 2.3 μm, were highly resistant to ultrasound. Only 7.5 and 7.7% of cell disruption were achieved for C. pyrenoidosa and M. aeruginosa FACHB 1343, respectively, when they were sonicated for 60 min. Declumping effect was dominant in these strains. This suggested that cellulose-rich-wall algal species might be much more resistant than hydroxyproline-rich glycoproteins, and peptidoglycan-wall species to sonication. It also revealed that the larger cell size was more susceptible to sonication the cell would be. This research provides useful insights into choosing the low-cost microalgae for extraction or controlling specific microalgal blooms in water systems using ultrasound.

ObjectivesKallikrein 7 (KLK7) is a secreted serine protease that plays important roles in skin desquamation and tumour progression, which makes it an attracting drug target. To guide the design of KLK7 inhibitors, a series of coumarin-based inhibitors were used to perform 3D-quantitative structure–activity relationship analysis.Methods3D conformations of 37 inhibitors were generated and used to construct CoMFA and CoMSIA models. Then a complex model between the inhibitors and KLK7 was built with molecular docking.Key findingsWith the training set, the CoMFA and CoMSIA models achieved q2 values of 0.521 and 0.498, and r2 values of 0.942 and 0.983, respectively. With the testing set, the predicted r2 values were 0.663 and 0.669, respectively, for CoMFA and CoMSIA. 3D contour maps from these two models identified steric and hydrophobic interactions as the most important molecular features of these inhibitors. Furthermore, molecular docking study was performed to understand the binding modes between these compounds and KLK7, in which the critical steric and hydrophobic interactions between the inhibitors and KLK7 were confirmed.ConclusionsSteric and hydrophobic interactions are critical in the efficient binding of KLK7 inhibitors. Our analysis would provide a meaningful guideline for the rational design of novel KLK7 inhibitors.