Wild-type p53 restores cell cycle control and inhibits gene amplification in cells with mutant p53 alleles.

Loss of cell cycle control and acquisition of chromosomal rearrangements such as gene amplification often occur during tumor progression, suggesting that they may be correlated. We show here that the wild-type p53 allele is lost when fibroblasts from patients with the Li-Fraumeni syndrome (LFS) are passaged in vitro. Normal and LFS cells containing wild-type p53 arrested in G1 when challenged with the uridine biosynthesis inhibitor PALA and did not undergo PALA-selected gene amplification. The converse occurred in cells lacking wild-type p53 expression. Expression of wild-type p53 in transformants of immortal and tumor cells containing mutant p53 alleles restored G1 control and reduced the frequency of gene amplification to undetectable levels. These studies reveal that p53 contributes to a metabolically regulated G1 check-point, and they provide a model for understanding how abnormal cell cycle progression leads to the genetic rearrangements involved in tumor progression.     

Purification and structural properties of gelsolin, a Ca2+-activated regulatory protein of macrophages

We describe the purification procedure and some of the physiochemical properties of gelsolin, a major Ca2+-dependent regulatory protein of actin gel-sol transformation in rabbit lung macrophages. Gelsolin accounts for the majority of Ca2+ control of actin gelation in macrophage extracts. It is a single polypeptide chain with an average molecular weight of 91,000 a Stokes radius of 44 A, a sedimentation coefficient (s20(0),w) of 4.9 S, an isoelectric point of 6.1, and a frictional ratio of 1.43. Gelsolin binds 2 mol of Ca2+ with high affinity (Ka 1.09 X 10(6) M-1) in the presence of 0.1 M KCl and 2 mM MgCl2.     

Reconstituted low density lipoprotein: A vehicle for the delivery of hydrophobic fluorescent probes to cells

Previous studies have shown that the cholesteryl ester core of plasma low density lipoprotein (LDL) can be extracted with heptane and replaced with a variety of hydrophobic molecules. In the present report we use this reconstitution technique to incorporate two fluorescent probes, 3-pyrenemethyl-23, 24-dinor-5-cholen-22-oate-3β-yl oleate (PMCA oleate) and dioleyl fluorescein, into heptane-extracted LDL. Both fluorescent lipoprotein preparations were shown to be useful probes for visualizing the receptor-mediated endocytosis of LDL in cultured human fibroblasts. When normal fibroblasts were incubated at 37°C with either of the fluorescent LDL preparations, fluorescent granules accumulated in the perinuclear region of the cell. In contrast, fibroblasts from patients with the homozygous form of familial hypercholesterolemia (FH) that lack functional LDL receptors did not accumulate visible fluorescent granules when incubated with the fluorescent reconstituted LDL. A fluorescence-activated cell sorter was used to quantify the fluorescence intensity of individual cells that had been incubated with LDL reconstituted with dioleyl fluorescein. With this technique a population of normal fibroblasts could be distinguished from a population of FH fibroblasts. The current studies demonstrate the feasibility of using fluorescent reconstituted LDL in conjunction with the cell sorter to isolate mutant cells lacking functional LDL receptors.     

piR-823 inhibits cell apoptosis via modulating mitophagy by binding to PINK1 in colorectal cancer

Mitophagy plays a vital role in the maintenance of mitochondrial homeostasis and tumorigenesis. Noncoding RNA piR-823 contributes to colorectal tumorigenesis. In this study, we aim to evaluate piR-823-mediated mitophagy and its mechanistic association with colorectal cancer (CRC). Digital gene expression analysis was performed to explore the potential functions of piR-823. A piR-823 antagomir (Ant-823) was used to inhibit piR-823 expression, and piR-823 mimics (mimics-823) were used to increase piR-823 expression. Mitophagy was measured in vivo and in vitro by immunofluorescence and western blot analysis. JC-1 staining, ATP production, real-time PCR, and western blot analysis were used to measure changes in mitochondrial quality and number. siRNA transfection was used to inhibit mitophagy, and CCCP was used to induce mitophagy. RNA pull-down assays and RNA-binding protein immunoprecipitation assays were conducted to investigate the molecular mechanisms. Here, we found that CRC cells transfected with Ant-823 presented an altered expression of autophagic and mitophagy genes by Digital gene expression analysis. Ant-823 could promote Parkin activation and mitophagy in vitro and in vivo, followed by mitochondrial loss and dysfunction of some mitochondria, whereas mimics-823 exerted the opposite effects in CRC cells. The inhibition of mitophagy by siParkin alleviated Ant-823-induced mitochondrial loss and dysfunction, as well as apoptosis to a certain extent. Furthermore, piR-823 was found to interact with PINK1 and promote its ubiquitination and proteasome-dependent degradation, thus alleviating mitophagy. Finally, these findings were verifed in samples obtained by patients affected by colorectal cancer. In conclusion, we identify a novel mechanism by which piR-823 regulates mitophagy during CRC tumorigenesis by increasing PINK1 degradation. Subject terms: Colorectal cancer, Gastrointestinal cancer     

Discovery of novel mRNA demethylase FTO inhibitors against esophageal cancer

A series of 1,2,3-triazole analogues as novel fat mass and obesity-associated protein (FTO) inhibitors were synthesised in this study. Among all 1,2,3-triazoles, compound C6 exhibited the most robust inhibition of FTO with an IC₅₀ value of 780 nM. It displayed the potent antiproliferative activity against KYSE-150, KYSE-270, TE-1, KYSE-510, and EC109 cell lines with IC₅₀ value of 2.17, 1.35, 0.95, 4.15, and 0.83 μM, respectively. In addition, C6 arrested the cell cycle at G2 phase against TE-1 and EC109 cells in a concentration-dependent manner. Analysis of cellular mechanisms demonstrated that C6 concentration-dependently regulated epithelial mesenchymal transition (EMT) pathway and PI3K/AKT pathway against TE-1 and EC109 cells. Molecular docking studies that C6 formed important hydrogen-bond interaction with Lys107, Asn110, Tyr108, and Leu109 of FTO. These findings suggested that C6 as a novel FTO inhibitor and orally antitumor agent deserves further investigation to treat esophageal cancer.     

A secreted ribonuclease effector from Verticillium dahliae localizes in the plant nucleus to modulate host immunity

The arms race between fungal pathogens and plant hosts involves recognition of fungal effectors to induce host immunity. Although various fungal effectors have been identified, the effector functions of ribonucleases are largely unknown. Herein, we identified a ribonuclease secreted by Verticillium dahliae (VdRTX1) that translocates into the plant nucleus to modulate immunity. The activity of VdRTX1 causes hypersensitive response (HR)‐related cell death in Nicotiana benthamiana and cotton. VdRTX1 possesses a signal peptide but is unlikely to be an apoplastic effector because its nuclear localization in the plant is necessary for cell death induction. Knockout of VdRTX1 significantly enhanced V. dahliae virulence on tobacco while V. dahliae employs the known suppressor VdCBM1 to escape the immunity induced by VdRTX1. VdRTX1 homologs are widely distributed in fungi but transient expression of 24 homologs from other fungi did not yield cell death induction, suggesting that this function is specific to the VdRTX1 in V. dahliae. Expression of site‐directed mutants of VdRTX1 in N. benthamiana leaves revealed conserved ligand‐binding sites that are important for VdRTX1 function in inducing cell death. Thus, VdRTX1 functions as a unique HR‐inducing effector in V. dahliae that contributes to the activation of plant immunity.     

长山群岛海域真核微藻粒级结构及扇贝摄食选择性

采用高通量测序-分子鉴定分级技术于2019年对长山群岛全海域真核微藻粒级结构进行了研究。结果发现,春季以中（47%）、小粒级（41%）为主,夏季以小（39%）、大粒级（38%）为主,秋季以大粒级（60%）为主,春、夏、秋季小、中、大粒级微藻比例为42：47：11、39：23：38、22：18：60。小粒级微藻优势种为细小微胞藻（Micromonas pusilla）、融合微胞藻（Micromonas commoda）和金牛微球藻（Ostreococcus tauri）,中粒级微藻优势种为剧毒卡尔藻（Karlodinium veneficum）、大粒级微藻优势种为柔弱几内亚藻（Guinardia delicatula）、平野亚历山大藻（Alexandrium hiranoi）、多纹膝沟藻（Gonyaulax polygramma）,综合整个真核微藻群落,春季由中粒径的剧毒卡尔藻占据优势（23.9%）,夏季由大粒径的平野亚历山大藻占据优势（29.4%）,秋季由大粒径的多纹膝沟藻占据优势（66.8%）,有毒甲藻在该海域中占有绝对优势,贝毒累积风险较高,小粒径微藻中金牛微球藻和抑食金球藻曾在渤海引发褐潮,潜在威胁该海域贝类养殖业。虾夷扇贝对小粒级和大粒级微藻的选择性较低,对中粒级微藻的选择性较高,尤其对水体中优势种剧毒卡尔藻一直表现出主动选择。光学需氧量、无机氮、溶解氧、石油类及部分重金属Cd、As、Hg影响着整个长山群岛海域真核微藻粒级结构时空演变。     

Caveolin-1 identified as a key mediator of acute lung injury using bioinformatics and functional research

Acute lung injury (ALI) is a potentially life-threatening, devastating disease with an extremely high rate of mortality. The underlying mechanism of ALI is currently unclear. In this study, we aimed to confirm the hub genes associated with ALI and explore their functions and molecular mechanisms using bioinformatics methods. Five microarray datasets available in GEO were used to perform Robust Rank Aggregation (RRA) to identify differentially expressed genes (DEGs) and the key genes were identified via the protein-protein interaction (PPI) network. Lipopolysaccharide intraperitoneal injection was administered to establish an ALI model. Overall, 40 robust DEGs, which are mainly involved in the inflammatory response, protein catabolic process, and NF-κB signaling pathway were identified. Among these DEGs, we identified two genes associated with ALI, of which the CAV-1/NF-κB axis was significantly upregulated in ALI, and was identified as one of the most effective targets for ALI prevention. Subsequently, the expression of CAV-1 was knocked down using AAV-shCAV-1 or CAV-1-siRNA to study its effect on the pathogenesis of ALI in vivo and in vitro. The results of this study indicated that CAV-1/NF-κB axis levels were elevated in vivo and in vitro, accompanied by an increase in lung inflammation and autophagy. The knockdown of CAV-1 may improve ALI. Mechanistically, inflammation was reduced mainly by decreasing the expression levels of CD3 and F4/80, and activating autophagy by inhibiting AKT/mTOR and promoting the AMPK signaling pathway. Taken together, this study provides crucial evidence that CAV-1 knockdown inhibits the occurrence of ALI, suggesting that the CAV-1/NF-κB axis may be a promising therapeutic target for ALI treatment.Subject terms: Cell signalling, Respiratory tract diseases