Treatment of neurologic pathology and inflammation in Machado-Joseph disease through in vivo self-assembled siRNA.

Machado-Joseph disease, also known as Spinocerebellar ataxia type 3 (MJD/SCA3), is a fatal autosomal dominant hereditary ataxia characterized by cerebellar ataxia resulting from the abnormal expansion of CAG repeats in exon 10 of the ATXN3 gene. Presently, there is no effective treatment for SCA3. Small interfering RNAs (siRNAs) are emerging as potential therapeutic strategies to specifically target the disease-causing mutant ATXN3 (mATXN3) protein. However, the delivery efficiency of siRNAs remains a major obstacle for clinical application, particularly in brain disorders. This study aimed to develop a synthetic biology strategy to reprogram the host liver as a tissue chassis to induce and deliver in vivo self-assembled siRNAs (IVSA-siRNAs) to target the ATXN3 gene. A synthetic construct directed by a cytomegalovirus promoter was designed to encode a neuron-targeting rabies virus glycoprotein tag and mATXN3-siRNA. After intravenous injection, the synthetic construct was taken up by mouse livers, which were then reprogrammed to enable the self-assembly, production, and secretion of small extracellular vesicles (sEVs) encapsulating mATXN3-siRNA. The sEV-encapsulated mATXN3-siRNA was further transported through the endogenous circulating system of sEVs, crossing the blood-brain barrier and reaching the cerebellar cortex and spinal cerebellar tract, where they silenced the ATXN3 gene. Treatment with the synthetic construct for 8 or 12 weeks led to significant improvements in motor balance ability and reduction of cerebellar atrophy in YACMJD84.2 transgenic mice. The number of Purkinje cells in the cerebellar cortex was significantly increased, and the loss of myelin basic protein was reduced. Moreover, the quantity of neurotoxic nuclear inclusion bodies and the expression of glial fibrillary acidic protein, which promotes neuroinflammation in activated astrocytes, were decreased significantly. The synthetic construct facilitated the generation and delivery of IVSA-siRNA to the cerebellar cortex and spinal cerebellar tract, thereby inhibiting the expression of mATXN3 protein. This treatment successfully addressed motor impairments, alleviated neuropathological phenotypes, and mitigated neuroinflammation in YACMJD84.2 transgenic mice. Our strategy effectively overcomes the primary challenges associated with siRNA therapy for cerebellar ataxia, offering a promising avenue for future clinical treatments.

Genome-wide identification and analysis of the cotton ALDH gene family.

BACKGROUND: Aldehyde dehydrogenases (ALDHs) are a family of enzymes that catalyze the oxidation of aldehyde molecules into the corresponding carboxylic acid, regulate the balance of aldehydes and protect plants from the poisoning caused by excessive accumulation of aldehydes; however, this gene family has rarely been studied in cotton. RESULTS: In the present study, genome-wide identification was performed, and a total of 114 ALDH family members were found in three cotton species, Gossypium hirsutum, Gossypium arboreum and Gossypium raimondii. The ALDH genes were divided into six subgroups by evolutionary analysis. ALDH genes in the same subgroup showed similar gene structures and conserved motifs, but some genes showed significant differences, which may result in functional differences. Chromosomal location analysis and selective pressure analysis revealed that the ALDH gene family had experienced many fragment duplication events. Cis-acting element analysis revealed that this gene family may be involved in the response to various biotic and abiotic stresses. The RT‒qPCR results showed that the expression levels of some members of this gene family were significantly increased under salt stress conditions. Gohir.A11G040800 and Gohir.D06G046200 were subjected to virus-induced gene silencing (VIGS) experiments, and the sensitivity of the silenced plants to salt stress was significantly greater than that of the negative control plants, suggesting that Gohir.A11G040800 and Gohir.D06G046200 may be involved in the response of cotton to salt stress. CONCLUSIONS: In total, 114 ALDH genes were identified in three Gossypium species by a series of bioinformatics analysis. Gene silencing of the ALDH genes of G. hirsutum revealed that ALDH plays an important role in the response of cotton to salt stress.

Interlocus Gene Conversion, Natural Selection, and Paralog Homogenization.

Following a duplication, the resulting paralogs tend to diverge. While mutation and natural selection can accelerate this process, they can also slow it. Here, we quantify the paralog homogenization that is caused by point mutations and interlocus gene conversion (IGC). Among 164 duplicated teleost genes, the median percentage of postduplication codon substitutions that arise from IGC rather than point mutation is estimated to be between 7% and 8%. By differentiating between the nonsynonymous codon substitutions that homogenize the protein sequences of paralogs and the nonhomogenizing nonsynonymous substitutions, we estimate the homogenizing nonsynonymous rates to be higher for 163 of the 164 teleost data sets as well as for all 14 data sets of duplicated yeast ribosomal protein-coding genes that we consider. For all 14 yeast data sets, the estimated homogenizing nonsynonymous rates exceed the synonymous rates.

A Novel Homozygous RHOH Variant Associated with T Cell Dysfunction and Recurrent Opportunistic Infections.

RHOH, an atypical small GTPase predominantly expressed in hematopoietic cells, plays a vital role in immune function. A deficiency in RHOH has been linked to epidermodysplasia verruciformis, lung disease, Burkitt lymphoma and T cell defects. Here, we report a novel germline homozygous RHOH c.245G > A (p.Cys82Tyr) variant in a 21-year-old male suffering from recurrent, invasive, opportunistic infections affecting the lungs, eyes, and brain. His sister also succumbed to a lung infection during early adulthood. The patient exhibited a persistent decrease in CD4+ T, B, and NK cell counts, and hypoimmunoglobulinemia. The patient's T cell showed impaired activation upon in vitro TCR stimulation. In Jurkat T cells transduced with RHOHC82Y, a similar reduction in activation marker CD69 up-regulation was observed. Furthermore, the C82Y variant showed reduced RHOH protein expression and impaired interaction with the TCR signaling molecule ZAP70. Together, these data suggest that the newly identified autosomal-recessive RHOH variant is associated with T cell dysfunction and recurrent opportunistic infections, functioning as a hypomorph by disrupting ZAP70-mediated TCR signaling.

DNA-Encoded Noncanonical Substrate Library for Protease Profiling.

Profiling the substrate sequence preferences of proteases is important for understanding both biological functions as well as for designing protease inhibitors. Several methods are available for profiling the sequence specificity of proteases. However, there is currently no rapid and high-throughput method to profile specificity of proteases for noncanonical substrates. In this study, we described a strategy to use a DNA-encoded noncanonical substrate library to identify the protease substrates composed of both canonical and noncanonical amino acids. This approach uses a DNA-encoded peptide library and introduces a biotin molecule at the N-terminus to immobilize the library on a solid support. Upon protease hydrolysis, the released DNA tag of the substrate peptides can be sequenced to identify the substrate structures. Using this approach, we profiled trypsin and fibroblast activation protein α and discovered noncanonical substrates that were more efficiently cleaved than the commonly used substrates. The identified substrates of FAP were further used to design corresponding covalent inhibitors containing non-canonical sequences with high potency for the target protease. Overall, our approach can aid in the development of new protease substrates and inhibitors.

Cytotoxic and viricidal effects of human semen on mumps virus-infected lymphocytes: In vitro studies.

Viruses in human semen may be sexually transmitted via free and cell-mediated viral infection. The potential effects of semen on the infection and sexual transmission of most viruses in semen remain largely unclear. The present study elucidated the inhibitory effects of human seminal plasma (SP) on Jurkat cell (JC)-mediated mumps virus (MuV) infection. We demonstrated that MuV efficiently infected JCs and that the JCs infected by MuV (JC-MuV) mediated MuV infection of HeLa cells. Remarkably, SP was highly cytotoxic to JCs and inhibited JC-MuV infection of HeLa cells. The cytotoxic factor possessed a molecular weight of less than 3 kDa, whereas that of the viricidal factor was over 100 kDa. The cooperation of cytotoxic and viricidal factors was required for the SP inhibition of JC-MuV infection, and prostatic fluid (PF) was responsible for both the cytotoxic and viricidal effects of SP. The cytotoxic effects we observed were resistant to the treatment of PF with boiling water, proteinase K, RNase A, and DNase I. Our results provide novel insights into the antiviral properties of SP, which may limit cell-mediated sexual viral transmission.

Glycosylation editing: an innovative therapeutic opportunity in precision oncology.

Cancer is still one of the most arduous challenges in the human society, even though humans have found many ways to try to conquer it. With our incremental understandings on the impact of sugar on human health, the clinical relevance of glycosylation has attracted our attention. The fact that altered glycosylation profiles reflect and define different health statuses provide novel opportunities for cancer diagnosis and therapeutics. By reviewing the mechanisms and critical enzymes involved in protein, lipid and glycosylation, as well as current use of glycosylation for cancer diagnosis and therapeutics, we identify the pivotal connection between glycosylation and cellular redox status and, correspondingly, propose the use of redox modulatory tools such as cold atmospheric plasma (CAP) in cancer control via glycosylation editing. This paper interrogates the clinical relevance of glycosylation on cancer and has the promise to provide new ideas for laboratory practice of cold atmospheric plasma (CAP) and precision oncology therapy.

Oxidation of Methanesulfinate by Hexachloroiridate(IV) and the Standard Electrode Potential of the Aqueous Methanesulfonyl Radical.

The aqueous reaction of [IrCl6]2- with CH3SO2- is biphasic and yields a 1:1 mixture of [IrCl6]3- and [IrCl5(H2O)]2- and CH3SO2Cl in the initial rapid phase. The next slow phase corresponds to the hydrolysis of CH3SO2Cl to yield CH3SO3- and Cl-. The initial phase shows kinetic inhibition by [IrCl6]3- that can be minimized by the addition of the radical scavenger propiolic acid. A detailed analysis of the kinetics indicates a mechanism with reversible outer-sphere electron transfer from CH3SO2- to [IrCl6]2- as the first step, followed by the irreversible inner-sphere oxidation of CH3SO2• by [IrCl6]2- to yield [IrCl5(H2O)]2- and CH3SO2Cl. Analysis of the inhibition by [IrCl6]3- and the kinetic effects of propiolic acid enable the determination of the equilibrium constant for the first electron-transfer step. This equilibrium constant then yields E° (CH3SO2•/CH3SO2-) = 1.01 V vs NHE at 25 °C. This is the first report of a standard potential for an alkanesulfonyl radical.

Myoglobin improves doxycycline sensitivity in pancreatic cancer through promoting heme oxygenase-1-mediated ferroptosis.

Ferroptosis is expected to be a therapeutic target for cancers including pancreatic cancer. We aimed to screen genes that regulate ferroptosis and doxycycline resistance in pancreatic cancer and to explore the underlying mechanisms. Bioinformatics analysis was performed to identify genes that respond to ferroptosis in two human pancreatic cancer cells with GOT1 knocked down or not. 325 and 842 genes were upregulated in MiaPaCa and Tu8902 cells in response to GOT1 knockdown, with 43 genes shared. Among the 43 genes, 14 genes were identified to interact with ferroptosis key genes. MB and HMOX1 were the genes most sensitive to Erastin and doxycycline. Moreover, MB and HMOX1 expression was higher in human normal pancreatic duct epithelial cells than in pancreatic cancer cells. MB and HMOX1 proteins physically bound and promoted each other's expression. By interacting with HMOX1, MB suppressed pancreatic cancer cell proliferation, colony formation and invasion, and promoted cell ferroptosis and sensitivity to erastin and doxycycline. Silencing HMOX1 reversed the promoting effect of MB on cell ferroptosis and sensitivity to doxycycline. A pancreatic cancer xenograft model was established by subcutaneous injection of Panc-1 cells transfected with or without Ad-MB, and doxycycline was administered intraperitoneally. Overexpression of MB enhanced the inhibitory effect of doxycycline on xenograft growth. In conclusion, MB facilitated doxycycline sensitivity in pancreatic cancer cells through promoting HMOX1-mediated ferroptosis.

Evaluation of net carbon sequestration and ecological benefits from single biochar-incorporated sorghum farmland systems in saline-alkali areas of Inner Mongolia, China.

Biochar is widely recognized as a soil amendment to reduce greenhouse gas emissions and enhance soil carbon storage in agroecosystems; however, the systematic focus on carbon balance and ecological benefits in cropping systems remains unclear in saline-alkali areas under water-saving irrigation. Here, a 2-yr field experiment with carbon footprint method was conducted to determine soil carbon budgets, biochar carbon efficiency performance, and the economic and ecological benefits of mulched drip-irrigated sorghum production, in an arid salinized region of Inner Mongolia, China. Corn straw-derived biochar dosages of 0 (CK), 15 (B15), 30 (B30), and 45 (B45) t hm-2 were just applied into the soil in the first crop growing season. A single application of biochar to soil significantly reduced CO2 emissions for the current and subsequent crop-growing seasons, with 13.1%, 16.7%, and 12.5% reductions for B15, B30, and B45, respectively. Compared with the non-biochar control plots, B15, B30, and B45 also increased NPP by 36.7%, 38.4%, and 27.1%, respectively. The actual effects on improving net carbon sequestration for B15, B30, and B45 in the first year were higher than those in the second year, with mean increases of 1.27, 1.47, and 1.36 times, respectively; however, the efficiencies of biochar for fixing carbon per biochar dosage input for B15 were 72.8% and 64.1% higher than those of B30 and B45, respectively. Net profits were significantly improved by 57.2-87.1% by biochar treatments. The environmental benefits of biochar carbon trading revenues for B15, B30, and B45 increased by 105.9%, 162.1%, and 109.6%, respectively. The minimum observation for carbon productivity and the maximum measurements for both the economic and ecological benefits were B15. The B15 also significantly increased sorghum yield and grain number. Results demonstrate that biochar application in the current growing season helps reduce soil carbon emissions, increases net carbon sequestration for current and subsequent sorghum agroecosystems, and enhances net profit and ecological benefits. The optimal positive synergistic effect was observed at a biochar application rate of 15 t hm-2 for reducing soil carbon emissions, increasing crop production, and improving the ecological environment.

Mechanism of an RBM-targeted rabbit monoclonal antibody 9H1 neutralizing SARS-CoV-2.

The COVID-19 pandemic, caused by SARS-CoV-2, has led to over 750 million infections and 6.8 million deaths worldwide since late 2019. Due to the continuous evolution of SARS-CoV-2, many significant variants have emerged, creating ongoing challenges to the prevention and treatment of the pandemic. Therefore, the study of antibody responses against SARS-CoV-2 is essential for the development of vaccines and therapeutics. Here we perform single particle cryo-electron microscopy (cryo-EM) structure determination of a rabbit monoclonal antibody (RmAb) 9H1 in complex with the SARS-CoV-2 wild-type (WT) spike trimer. Our structural analysis shows that 9H1 interacts with the receptor-binding motif (RBM) region of the receptor-binding domain (RBD) on the spike protein and by directly competing with angiotensin-converting enzyme 2 (ACE2), it blocks the binding of the virus to the receptor and achieves neutralization. Our findings suggest that utilizing rabbit-derived mAbs provides valuable insights into the molecular interactions between neutralizing antibodies and spike proteins and may also facilitate the development of therapeutic antibodies and expand the antibody library.

Modular Click Assembly DNA-Encoded Glycoconjugate Libraries with on-DNA Functional Group Transformations.

Carbohydrates are an important class of naturally active products and play vital roles in regulating various physiological activities. To meet the demand for carbohydrate-based libraries used for the identification of potential drug candidates for pharmaceutical-related targets, we developed a set of on-DNA protocols to construct the DNA-encoded glycoconjugates, including Seyferth-Gilbert homologation, anomeric azidation, and CuAAC cyclization. These on-DNA chemistries enable the generation and modification of DNA-linked glycosyl compounds with good conversions and broad substrate scope. Finally, three DNA-linked glycoconjugate libraries were successfully generated to demonstrate their applicability and feasibility in library preparation.

Immunological effects of recombinant Lactobacillus casei expressing pilin MshB fused with cholera toxin B subunit adjuvant as an oral vaccine against Aeromonas veronii infection in crucian carp.

Aeromonas veronii is a zoonotic agent capable of infecting fish and mammals, including humans, posing a serious threat to the development of aquaculture and public health safety. Currently, few effective vaccines are available through convenient routes against A. veronii infection. Herein, we developed vaccine candidates by inserting MSH type VI pili B (MshB) from A. veronii as an antigen and cholera toxin B subunit (CTB) as a molecular adjuvant into Lactobacillus casei and evaluated their immunological effect as vaccines in a crucian carp (Carassius auratus) model. The results suggested that recombinant L. casei Lc-pPG-MshB and Lc-pPG-MshB-CTB can be stably inherited for more than 50 generations. Oral administration of recombinant L. casei vaccine candidates stimulated the production of high levels of serum-specific immunoglobulin M (IgM) and increased the activity of acid phosphatase (ACP), alkaline phosphatase (AKP) superoxide dismutase (SOD), lysozyme (LZM), complement 3 (C3) and C4 in crucian carp compared to the control group (Lc-pPG612 group and PBS group) without significant changes. Moreover, the expression levels of interleukin-10 (IL-10), interleukin-1ß (IL-1ß), tumour necrosis factor-α (TNF-α) and transforming growth factor-ß (TGF-ß) genes in the gills, liver, spleen, kidney and gut of crucian carp orally immunized with recombinant L. casei were significantly upregulated compared to the control groups, indicating that recombinant L. casei induced a significant cellular immune response. In addition, viable recombinant L. casei can be detected and stably colonized in the intestine tract of crucian carp. Particularly, crucian carp immunized orally with Lc-pPG-MshB and Lc-pPG-MshB-CTB exhibited higher survival rates (48% for Lc-pPG-MshB and 60% for Lc-pPG-MshB-CTB) and significantly reduced loads of A. veronii in the major immune organs after A. veronii challenge. Our findings indicated that both recombinant L. casei strains provide favorable immune protection, with Lc-pPG-MshB-CTB in particular being more effective and promising as an ideal candidate for oral vaccination.

Circ_0058063 regulates cell vitality and proliferation in oesophageal squamous-cell carcinomas.

Oesophageal squamous-cell carcinoma (ESCC) is a malignant tumor of the digestive system with a poor prognosis. Recent studies have shown the promoting effect of hsa_circ_0058063 (circ_0058063) on ESCC, but the potential regulatory mechanisms of circ_0058063 in ESCC remain largely unclear. The levels of circ_0058063, microRNA-4319 (miR-4319) and mRNA of thrombospondin-1 (THBS1) were indicated by quantitative real-time polymerase chain reaction in ESCC tissues and cells. Meanwhile, the level of THBS1 was quantified by western blot analysis. In addition, the cell functions were examined by CCK8 assay, Edu assay, flow cytometry assay and transwell assay. Furthermore, the interplay between miR-4319 and circ_0058063 or THBS1 was detected by dual-luciferase reporter assay. Finally, an in vivo experiment was implemented to confirm the effect of circ_0058063. The level of circ_0058063 and THBS1 were increased, and the miR-4319 level was decreased in ESCC tissues in contrast to that in normal tissues and cells. For functional analysis, circ_0058063 deficiency inhibited cell vitality, cell proliferation, migration and invasion in ESCC cells, whereas promoted cell apoptosis. Moreover, miR-4319 was confirmed to repress the progression of ESCC cells by suppressing THBS1. In mechanism, circ_0058063 acted as a miR-4319 sponge to regulate the level of THBS1. Besides, circ_0058063 knockdown also attenuated tumour growth in vivo. Circ_0058063 facilitates the development of ESCC through increasing THBS1 expression by regulating miR-4319, which also offered an underlying targeted therapy for ESCC treatment.

Inferring single cell expression profiles from overlapped pooling sequencing data with compressed sensing strategy.

Though single cell RNA sequencing (scRNA-seq) technologies have been well developed, the acquisition of large-scale single cell expression data may still lead to high costs. Single cell expression profile has its inherent sparse properties, which makes it compressible, thus providing opportunities for solutions. Here, by computational simulation as well as experiment of 54 single cells, we propose that expression profiles can be compressed from the dimension of samples by overlapped assigning each cell into plenty of pools. And we prove that expression profiles can be inferred from these pool expression data with overlapped pooling design and compressed sensing strategy. We also show that by combining this approach with plate-based scRNA-seq measurement, it can maintain its superiorities in gene detection sensitivity and individual identity and recover the expression profile with high precision, while saving about half of the library cost. This method can inspire novel conceptions on the measurement, storage or computation improvements for other compressible signals in many biological areas.

Ultra-sensitive detection of tumor necrosis factor alpha based on silver-coated gold core shell and magnetically separated recognition of SERS aptamer sensors.

A highly sensitive tumor necrosis factor α (TNF-α) detection method based on a surface-enhanced Raman scattering (SERS) magnetic patch sensor is reported. Magnetic beads (MNPs) and core shells were used as the capture matrix and signaling probe, respectively. For this purpose, antibodies were immobilized on the surface of magnetic beads, and then Au@4-MBN@Ag core-shell structures coupled with aptamers and TNF-α antigen were added sequentially to form a sandwich immune complex. Quantitative analysis was performed by monitoring changes in the characteristic SERS signal intensity of the Raman reporter molecule 4-MBN. The results showed that the limit of detection (LOD) of the proposed method was 4.37 × 10-15 mg·mL-1 with good linearity (R2 = 0.9918) over the concentration range 10-12 to 10-5 mg·mL-1. Excellent assay accuracy was also demonstrated, with recoveries in the range 102% to 114%. Since all reactions occur in solution and are separated by magnetic adsorption of magnetic beads, this SERS-based immunoassay technique solves the kinetic problems of limited diffusion and difficult separation on solid substrates. The method is therefore expected to be a good clinical tool for the diagnosis of the inflammatory biomarker THF-α and in vivo inflammation screening.

Identifying priority areas for freshwater supply conservation integrating multi-scale freshwater flows.

Identifying priority areas for conservation is an effective measurement for the sustainable provision of ecosystem services (ESs) under threats globally. Although many approaches have been developed to identify conservation priority areas by combining supply and demand of ESs, the integration of ESs flows into the identification still need further exploration. For ESs like freshwater supply services, the processes of freshwater flows across multiple scales are crucial. This study aimed to propose a new study framework to identify priority areas for freshwater supply conservation by integrating the multi-scale (i.e., sub-watershed, tributary, and mainstream) freshwater flows, using the Yangtze River Delta as the study area. The results suggested that spatial mismatches between the supply and demand of freshwater supply services existed at different scales. There were approximately 129, 58, and 55 pairs of freshwater flows in sub-watersheds, tributaries, and mainstreams, respectively, which transported 5.98 × 1010 m3, 2.07 × 1010 m3 and 2.50 × 1010 m3 of freshwater. The results of multi-scale freshwater flows were integrated into conservation target goals, and the identified priority areas for freshwater supply conservation were selected at three scales. The priority areas selected at the sub-watershed scale were the largest. Compared with the traditional method of identifying priority areas without considering freshwater flows, the priority areas identified in this study included both sites with a high supply capacity and sites with a relatively low supply capacity, as they were significant for meeting the local freshwater demand. The increasing understanding of freshwater flows and the integration of the flows for the identification of priority areas for freshwater supply conservation are important for the development of more practical and rational policies or ecological management for the sustainable conservation of ESs.

Characterization of copper binding to different molecular weight fractions of dissolved organic matter in surface water.

Dissolved organic matter (DOM) is a kind of substance with complex compositions and wide molecular weight distribution, which can strongly combine with various pollutants. Therefore, the binding characteristics of DOM and heavy metal pollutants can be studied specifically according to the binding characteristics of DOM and pollutants. In this study, DOM in surface water bodies was divided into three levels (MW < 1 kDa, 1 kDa < MW < 5 kDa, MW > 5 kDa) according to different molecular weights (MW). The binding properties were investigated by fluorescence spectrum analysis and complex model. Four components (C1-C4) were identified by PARAFAC. Among them, the contribution rate of protein-like components C1, C2 and C4 to the total fluorescence intensity reached more than 78%, and the log Ka values of low molecular weight components were the highest, which were 3.28, 3.14 and 3.47, respectively, indicating higher binding ability with Cu2+.With the decrease of molecular weight, the log Kb value increases, indicating that the complexation is more stable. The humic component C3 in high molecular weight has stronger binding stability with Cu2+, but the number of binding sites for C3 is 0.36, while that for C2 is 1.51, indicating that its binding sites and binding ability are relatively low. The results showed that the DOM ligand of Cu2+ in surface water showed a certain molecular weight dependence. In addition, different MW DOM lead to different pollution forms. Different properties of DOM ligand combined with Cu2+ were studied in order to control the migration, transformation, bioavailability, morphology and stability of heavy metal pollutants, and to provide theoretical support for the practical application management of surface water pollution control.

WBP2 promotes gastric cancer cell migration via novel targeting of LATS2 kinase in the Hippo tumor suppressor pathway.

Dysregulation of signaling pathways is responsible for many human diseases. The lack of understanding of the molecular etiology of gastric cancer (GC) poses a substantial challenge to the development of effective cancer therapy. To better understand the molecular mechanisms underlying the pathogenesis of GC, which will facilitate the identification and development of effective therapeutic approaches to improve patient outcomes, mass spectrometry-based phosphoproteomics analysis was performed to map the global molecular changes in GC. A total of 530 proteins with altered phosphorylation levels were detected across a panel of 15 normal and GC cell lines. WW domain-binding protein 2 (WBP2) was validated to be upregulated in a subset of GC cell lines. WBP2 is overexpressed in 61% cases of GC compared to non-cancer tissues and high WBP2 expression correlates with poor clinical outcomes. WBP2 was found to be required for GC cell migration but is dispensable for cell growth and proliferation. WBP2 knockdown increased p-LATS2 with a concomitant increase in p-YAP, resulting in the cytoplasmic retention of YAP and ultimately the inhibition of YAP/TEAD activity and downregulation of TEAD target genes--CTGF and CYR61. Importantly, the loss of LATS2 reversed the activation of Hippo pathway caused by WBP2 knockdown, indicating that WBP2 acts through LATS2 to exert its function on the Hippo pathway. Moreover, WBP2 interacted with LATS2 to inhibit its phosphorylation and activity. In conclusion, our study established a pivotal role for WBP2 in the promotion of GC cell migration via a novel mechanism that inactivates the Hippo pathway transducer LATS2.

Molecular mechanism of albumin in suppressing invasion and metastasis of hepatocellular carcinoma.

BACKGROUND & AIMS: Worldwide, hepatocellular carcinoma (HCC) is one of the most common causes of death in people. Albumin (ALB) is considered as an important indicator for HCC prognosis, and evidence has shown HCC cell growth can be regulated by ALB. However, the role of ALB in hepatocarcinogenesis and the mechanism of action is still unknown. METHODS: The expression of ALB was determined by clinical profiles, immunohistochemistry, and western blot. Wound healing and Transwell assays were conducted to evaluate the effects of ALB during migration and invasion in HCC. We used mass spectrometry coupled isobaric tags for relative and absolute quantitation (iTRAQ)-technology to identify secretory differentially expressed proteins (DEPs) in ALB knockdown HepG2 cells. Western blot, reverse transcription-quantitative polymerase chain reaction and enzyme-linked immunosorbent assay techniques were used for verification. RESULTS: We suggested that ALB was associated with aggressive metastasis and depleting ALB significantly promoted invasion and migration of HCC. A total of 210 DEPs were identified after silencing of ALB. We observed that a negative correlation between ALB and urokinase plasminogen activator surface receptor (uPAR) expression levels. CONCLUSIONS: ALB acts as a tumour suppressor and plays a key role in HCC progression, particularly in invasion and metastasis. Suppression of ALB promoted migration and invasion of HCC cells by increasing uPAR, matrix metalloproteinase (MMP2), and MMP9.