Surface-enhanced resonance Raman spectroscopy and spectroscopy study of redox-induced conformational equilibrium of cytochrome c adsorbed on DNA-modified metal electrode

The redox-induced conformational equilibrium of cytochrome c (cyt c) adsorbed on DNA-modified metal electrode and the interaction mechanism of DNA with cyt c have been studied by electrochemical, spectroscopic and spectroelectrochemical techniques. The results indicate that the external electric field induces potential-dependent coordination equilibrium of the adsorbed cyt c between its oxidized state (with native six-coordinate low-spin and non-native five-coordinate high-spin heme configuration) and its reduced state (with native six-coordinate low-spin heme configuration) on DNA-modified metal electrode. The strong interactions between DNA and cyt c induce the self-aggregation of cyt c adsorbed on DNA. The orientational distribution of cyt c adsorbed on DNA-modified metal electrode is potential-dependent, which results in the deviation from an ideal Nernstian behavior of the adsorbed cyt c at high electrode potentials. The electric-field-induced increase in the activation barrier of proton-transfer steps attributed to the rearrangement of the hydrogen bond network and the self-aggregation of cyt c upon adsorption on DNA-modified electrode strongly decrease the interfacial electron transfer rate. In addition, the strongly Coulombic interactions between DNA and cyt c only disturb the microenvironment of the heme, and do not affect the states of heme ligation and spin. The secondary structure of the adsorbed cyt c is retained, while the conformation of DNA is changed from the B form DNA to A form DNA.     

HIV-1 glycoprotein 41 ectodomain induces activation of the CD74 protein-mediated extracellular signal-regulated kinase/mitogen-activated protein kinase pathway to enhance viral infection

Besides mediating the viral entry process, the human immunodeficiency virus (HIV-1) envelope protein gp41 can bind to many host cell components and regulate cell functions. Using a yeast two-hybrid system, we screened a human bone marrow cDNA library and identified a novel gp41-binding protein, CD74 (the MHC class II-associated invariant chain). Here, we report possible biological effects mediated by interaction between gp41 and CD74. We found that HIV-1 gp41 could bind directly to host CD74 in HIV-1-infected cells, and the peptide 6358 derived from gp41 loop region (aa 597-611) could effectively block the gp41-CD74 interaction. As a result of this binding, recombinant soluble gp41 and gp41 peptide 6358 activated the CD74-mediated ERK/MAPK pathway and significantly enhanced HIV-1 infection in vitro. Conversely, the enhancing effect could be suppressed by the recombinant CD74 extracellular domain. These results reveal a novel mechanism underlying gp41 mediation of HIV-1 infection and replication.     

MiR‐616‐3p modulates cell proliferation and migration through targeting tissue factor pathway inhibitor 2 in preeclampsia

Objectives

Despite improvements in diagnosis and treatment, preeclampsia (PE) continues to pose a significant risk of maternal and foetal morbidity and mortality if not addressed promptly. An increasing number of studies have suggested that tissue factor pathway inhibitor 2 (TFPI2) acts as a suppressor gene, possibly inhibiting multiple serine proteases affecting cell proliferation and migration. It plays an essential role in the occurrence and development of PE, but the pathogenesis remains unclear.

Materials and methods

In our research, we performed western blotting, immunohistochemistry and qPCR assays to investigate TFPI2 and miR‐616‐3p expression in preeclamptic placental tissues. Cell assays were performed in HTR‐8/SVneo and JEG3 cell lines. Cell proliferation and migration events were investigated by MTT, EdU and transwell assays. In conjunction with bioinformatics analysis, luciferase reporter assays were performed to elucidate the mechanism by which miR‐616‐3p binds to TFPI2 mRNA.

Results

We established that TFPI2 protein levels were significantly upregulated in PE placental tissues. In addition, we found that miR‐616‐3p binds specifically to the 3′‐UTR region of TFPI2 mRNA. Furthermore, miR‐616‐3p knockdown or TFPI2 overexpression substantially impaired cell growth and migration, whereas miR‐616‐3p upregulation or TFPI2 knockdown stimulated cell proliferation and migration. This miR‐616‐3p / TFPI2 axis was also found to affect the epithelial‐mesenchymal transition process in PE.

Conclusions

Our results demonstrated that TFPI2 plays a vital role in the progression of PE and might provide a prospective therapeutic strategy to mitigate the severity of the disorder.

     

错配修复基因与涎腺肿瘤的关系

错配修复(mismatch repair,MMR)是DNA复制后的一种修复机制,对维持基因组稳定起重要作用.错配修复基因功能缺陷是继癌基因激活、抑癌基因失活之后又一肿瘤的发生、发展机制,错配修复基因的异常表达与全身多种肿瘤相关.涎腺肿瘤为口腔颌面部常见肿瘤之一,具有与其他系统肿瘤相似的组织学类型,多来源于肌上皮.近年来,有关涎腺肿瘤与错配修复基因的关系正逐步成为研究热点,本文就错配修复基因的组成、作用机制以及与涎腺肿瘤发生、发展的关系作一综述.     

Antiviral flavonoids from the root bark of Morus alba L

A prenylated flavonoid, moralbanone, along with seven known compounds kuwanon S, mulberroside C, cyclomorusin, eudraflavone B hydroperoxide, oxydihydromorusin, leachianone G and alpha-acetyl-amyrin were isolated from the root bark of Morus alba L. Leachianone G showed potent antiviral activity (IC(50) = 1.6 microg/ml), whereas mulberroside C showed weak activity (IC(50) = 75.4 microg/ml) against herpes simplex type 1 virus (HSV-1). Their structures were elucidated by spectroscopic methods.     

Endotoxin/lipopolysaccharide activates NF-kappa B and enhances tumor cell adhesion and invasion through a beta 1 integrin-dependent mechanism

Beta(1) integrins play a crucial role in supporting tumor cell attachment to and invasion into the extracellular matrix. Endotoxin/LPS introduced by surgery has been shown to enhance tumor metastasis in a murine model. Here we show the direct effect of LPS on tumor cell adhesion and invasion in extracellular matrix proteins through a beta(1) integrin-dependent pathway. The human colorectal tumor cell lines SW480 and SW620 constitutively expressed high levels of the beta(1) subunit, whereas various low levels of alpha(1), alpha(2), alpha(4), and alpha(6) expression were detected. SW480 and SW620 did not express membrane-bound CD14; however, LPS in the presence of soluble CD14 (sCD14) significantly up-regulated beta(1) integrin expression; enhanced tumor cell attachment to fibronectin, collagen I, and laminin; and strongly promoted tumor cell invasion through the Matrigel. Anti-beta(1) blocking mAbs (4B4 and 6S6) abrogated LPS- plus sCD14-induced tumor cell adhesion and invasion. Furthermore, LPS, when combined with sCD14, resulted in NF-kappaB activation in both SW480 and SW620 cells. Inhibition of the NF-kappaB pathway significantly attenuated LPS-induced up-regulation of beta(1) integrin expression and prevented tumor cell adhesion and invasion. These results provide direct evidence that although SW480 and SW620 cells do not express membrane-bound CD14, LPS in the presence of sCD14 can activate NF-kappaB, up-regulate beta(1) integrin expression, and subsequently promote tumor cell adhesion and invasion. Moreover, LPS-induced tumor cell attachment to and invasion through extracellular matrix proteins is beta(1) subunit-dependent.     

长效促胰岛素降糖酵母的构建及其对糖尿病模型小鼠的治疗效果

益生菌生物药物是指通过口服表达药用多肽(蛋白)的重组益生菌活细胞达到治疗疾病的新型口服给药系统。为了构建一种能有效防治2型糖尿病的酵母生物药物,文章首先构建了酿酒酵母(S.cerevisiae)整合型表达载体pNK1-PGK,并且通过绿色荧光蛋白(GFP)证明其表达功能正常,利用该载体将10×GLP-1 (Glucagon-like peptide-1)基因转化到酿酒酵母INVSc1中,通过营养缺陷型和Western blotting成功筛选出表达10×GLP-1的长效促胰岛素降糖酵母(Long-acting GLP-1 hypoglycemic yeast, LHY)。该酵母生长迅速,外源基因10×GLP-1表达稳定,表达量达到1.56 mg/g细胞湿重。通过链脲佐菌素和高脂高糖饮食联合诱导的方法构建了2型糖尿病小鼠模型,用LHY对其进行口服灌胃治疗,证明LHY具有较好疗效,明显降低血糖水平。     

Effects of Cadmium on Root Growth, Cell Division and Nucleoli in Root Tip Cells of Garlic

The effects of different concentrations (10⁻⁷ to 10⁻² M) of cadmium chloride on root growth, cell division and nucleoli in root tip cells of Allium sativum L. were investigated. At lower concentrations of Cd²⁺ (10⁻⁷ to 10⁻⁶ M), Cd²⁺ did not influence the root growth, even had a stimulation effects during a short treatment. The results showed that the rate of root growth per day at the treatment groups (10⁻⁴ to 10⁻² M Cd²⁺) decreased with increasing duration of the treatment and increasing Cd²⁺ concentration. Cd²⁺ induced c-mitosis, anaphase bridges, chromosome stickiness and on nucleoli, causing some particles of similar silver-stained material scattered in the nuclei and making the silver staining reaction at the periphery of the nucleolus weaker. This revised version was published online in July 2006 with corrections to the Cover Date.     

Gating of inward rectifier K+ channels by proton-mediated interactions of N- and C-terminal domains

Ion channels play an important role in cellular functions, and specific cellular activity can be produced by gating them. One important gating mechanism is produced by intra- or extracellular ligands. Although the ligand-mediated channel gating is an important cellular process, the relationship between ligand binding and channel gating is not well understood. It is possible that ligands are involved in the interactions of different protein domains of the channel leading to opening or closing. To test this hypothesis, we studied the gating of Kir2.3 (HIR) by intracellular protons. Our results showed that hypercapnia or intracellular acidification strongly inhibited these channels. This effect relied on both the N and C termini. The CO(2)/pH sensitivities were abolished or compromised when one of the intracellular termini was replaced. Using purified N- and C-terminal peptides, we found that the N and C termini bound to each other in vitro. Although their binding was weak at pH 7.4, stronger binding was seen at pH 6.6. Two short sequences in the N and C termini were found to be critical for the N/C-terminal interaction. Interestingly, there was no titratable residue in these motifs. To identify the potential protonation sites, we systematically mutated most histidine residues in the intracellular N and C termini. We found that mutations of several histidine residues in the C but not the N terminus had a major effect on channel sensitivities to CO(2) and pH(i). These results suggest that at acidic pH, protons appear to interact with the C-terminal histidine residues and present the C terminus to the N terminus. Consequentially, these two intracellular termini bound to each other through two short motifs and closed the channel. Thus, a novel mechanism for K(+) channel gating is demonstrated, which involves the N- and C-terminal interaction with protons as the mediator.