ZNF403,一个新的细胞周期调节因子的功能研究(英文)

ZNF403和LCRG1是人类基因ZNF403的2个不同转录剪切本.以往的研究表明LCRG1在喉癌细胞株Hep-2中具有抑瘤特性.本研究旨在探明ZNF403和LCRG1不同剪切本之间的关系以及在肿瘤细胞中对ZNF403的功能进行研究.首先,采用实时荧光定量PCR对这2个转录本的相对表达水平进行分析,结果表明,ZNF403表达水平在不同细胞株中明显高于LCRG1(>10倍),为该基因的主要转录表达产物.随后分别采用MTT细胞生长分析法和裸鼠体内成瘤实验在体外和体内对ZNF403的功能进行分析,结果显示ZNF403的基因沉默可以同时在体内和体外抑制喉癌细胞Hep-2细胞的生长.为了探明其作用机制,本研究还采用细胞信息学、流式细胞周期分析术和高通量PCR点阵分析方法进一步分析,结果表明,ZNF403的基因沉默可显著抑制细胞DNA的复制并延缓细胞周期进入到有丝分裂期.同时发现ZNF403可调节一系列的细胞周期调节蛋白如MCM2、p21、ATM、MRE11A等.综上研究提示ZNF403为一新的细胞周期调节因子,其功能的缺失与肿瘤发生发展密切相关.

IL-17RD （Sef or IL-17RLM） interacts with IL-17 receptor and mediates IL-17 signaling

Interleukin-17 （IL-17 or IL-17A） production is a hallmark of TH17 cells, a new unique lineage of CD4^＋ T lymphocytes contributing to the pathogenesis of multiple autoimmune and inflammatory diseases. IL-17 receptor （IL-17R or IL-17RA） is essential for IL-17 biological activity. Emerging data suggest that the formation of a heteromeric and/or homomeric receptor complex is required for IL-17 signaling. Here we show that the orphan receptor IL-17RD （Sef, similar expression to FGF genes or IL-17RLM） is associated and colocalized with IL-17R. Importantly, IL-17RD mediates IL-17 signaling, as evaluated using a luciferase reporter driven by the native promoter of 24p3, an IL-17 target gene. In addition, an IL-17RD mutant lacking the intraeellnlar domain dominant-negatively suppresses IL-17R- mediated IL-17 signaling. Moreover, IL-17RD as well as IL-17R is associated with TRAF6, an IL-17R downstream molecule. These results indicate that IL-17RD is a part of the IL-17 receptor signaling complex, therefore providing novel evidence for IL-17 signaling through a heteromeric and/or homomeric receptor complex.

Regulation of epithelium-specific Ets-like factors ESE-1 and ESE-3 in airway epithelial cells： potential roles in airway inflammation

Airway inflammation is the hallmark of many respiratory disorders, such as asthma and cystic fibrosis. Changes in airway gene expression triggered by inflammation play a key role in the pathogenesis of these diseases. Genetic linkage studies suggest that ESE-2 and ESE-3, which encode epithelium-specific Ets-domain-containing transcription factors, are candidate asthma susceptibility genes. We report here that the expression of another member of the Ets family transcription factors ESE-1, as well as ESE-3, is upregulated by the inflammatory cytokines interleukin-1β （IL-1β） and tumor necrosis factor-α （TNF-α） in bronchial epithelial cell lines. Treatment of these cells with IL-1β and TNF-α resulted in a dramatic increase in mRNA expression for both ESE-1 and ESE-3. We demonstrate that the induced expression is mediated by activation of the transcription factor NF-κB. We have characterized the ESE-1 and ESE-3 promoters and have identified the NF-κB binding sequences that are required for the cytokine-induced expression. In addition, we also demonstrate that ESE-1 upregulates ESE-3 expression and downregulates its own induction by cytokines. Finally, we have shown that in E/f3 （homologous to human ESE-1） knockout mice, the expression of the inflammatory cytokine interleukin-6 （IL-6） is downregulated. Our findings suggest that ESE-1 and ESE-3 play an important role in airway inflammation.

Epithelium-specific ets transcription factor 2 upregulates cytokeratin 18 expression in pulmonary epithelial cells through an interaction with cytokeratin 18 intron 1

The role of Ese-2,an Ets family transcription factor,in gene regulation is not known.In this study,the interactionbetween Ese-2 and cytokeratin 18(K18)intron 1 was characterized in lung epithelial cells.Reporter gene assays showedEse-2 was able to upregulate K18 intron 1 enhanced reporter gene expression by approximately 2-fold.We found thatfull length Ese-2 did not bind DNA strongly,therefore truncated versions of the protein,containing the ETS domain orPointed domain,were created and tested in electrophoresis mobility shift assays.Multiple interactions between the ETSdomain and putative DNA binding sites within K18 intron 1 were observed,which led to the determination of a possibleEse-2 DNA binding consensus sequence.These experiments suggest that Ese-2 could play a role in the regulation ofK18 expression in lung epithelial cells.

“注册使用人数”有价值吗?

入门网站和其他网际网路业者总是吹嘘他们的注册使用人数,甚至把这些数字纳入每季业绩报告中,作为成功的一种衡量标准。但分析师质疑,注册使用人数似嫌夸大,且各家使用者多有重叠,除粉饰业绩报告外,尚未能转化为获利。

For your eyes only:Harnessing human embryonic stem cell-derived retinal pigment epithelial cells to improve impaired vision

Vision loss or impairment resulting from the degeneration of the retinal pigment epithelium and photoreceptor death affects millions worldwide.Recent exciting results from clinical studies of small numbers of patients treated with human embryonic stem cell-derived retinal pigment epithelial cells may provide hope for affected individuals.

The key piece of the airway remodeling puzzle revealed-LIGHT signaling in the lung

Asthma is a chronic lung disease that primarily affects the lower respiratory tract in more than 300 million people worldwide.Asthma is often triggered by exposure to certain substances or conditions,such as allergens or cold air.The overreaction to the allergens causes inflammation and narrowing of the airway,which leads to episodic or persistent symptoms including shortness of breath,nocturnal cough,chest tightness,and wheezing.

Gene therapy: light is finally in the tunnel

After two decades of ups and downs,gene therapy has recently achieved a milestone in treating patients with Leber’s congenital amaurosis(LCA).LCA is a group of inherited blinding diseases with retinal degeneration and severe vision loss in early infancy.Mutations in several genes,including RPE65,cause the disease.Using adenoassociated virus as a vector,three independent teams of investigators have recently shown that RPE65 can be delivered to retinal pigment epithelial cells of LCA patients by subretinal injections resulting in clinical benefits without side effects.However,considering the whole field of gene therapy,there are still major obstacles to clinical applications for other diseases.These obstacles include innate and immune barriers to vector delivery,toxicity of vectors and the lack of sustained therapeutic gene expression.Therefore,new strategies are needed to overcome these hurdles for achieving safe and effective gene therapy.In this article,we shall review the major advancements over the past two decades and,using lung gene therapy as an example,discuss the current obstacles and possible solutions to provide a roadmap for future gene therapy research.

Potential of Helper-Dependent Adenoviral Vectors in Modulating Airway Innate Immunity

Innate immune responses form the first line of defense against foreign insults and recently significant advances have been made in our understanding of the initiation of innate immune response along with its ability to modulate inflammation. In airway diseases such as asthma, COPD and cystic fibrosis, over reacting of the airway innate immune responses leads to cytokine imbalance and airway remodeling or damage. Helper-dependent adenoviral vectors have the potential to deliver genes to modulate airway innate immune responses and have many advantages over its predecessors. However, there still are a few limitations that need to be addressed prior to their use in clinical applications.

Lung gene therapydHow to capture illumination from the light already present in the tunnel

Gene therapy has been considered as the most ideal medical intervention for genetic diseases because it is intended to target the cause of diseases instead of disease symptoms.Availability of techniques for identification of genetic mutations and for in vitro manipulation of genes makes it practical and attractive.After the initial hype in 1990s and later disappointments in clinical trials formore than a decade,light has finally come into the tunnel in recent years,especially in the field of eye gene therapy where it has taken big strides.Clinical trials in gene therapy for retinal degenerative diseases such as Leber’s congenital amaurosis(LCA)and choroideremia demonstrated clear therapeutic efficacies without apparent side effects.Although these successful examples are still rare and sporadic in the field,they provide the proof of concept for harnessing the power of gene therapy to treat genetic diseases and to modernize our medication.In addition,those success stories illuminate the path for the development of gene therapy treating other genetic diseases.Because of the differences in target organs and cells,distinct barriers to gene delivery exist in gene therapy for each genetic disease.It is not feasible for authors to review the current development in the entire field.Thus,in this article,we will focus onwhatwe can learn from the current success in gene therapy for retinal degenerative diseases to speed up the gene therapy development for lung diseases,such as cystic fibrosis.

Highly efficient retinal gene delivery with helper-dependent adenoviral vectors

There have been significant advancements in the field of retinal gene therapy in the past several years.In particular,therapeutic efficacy has been achieved in three separate human clinical trials conducted to assess the ability of adeno-associated viruses(AAV)to treat of a type of Leber’s congenital amaurosis caused by RPE65 mutations.However,despite the success of retinal gene therapy with AAV,challenges remain for delivering large therapeutic genes or genes requiring long DNA regulatory elements for controlling their expression.For example,Stargardt’s disease,a form of juvenile macular degeneration,is caused by defects in ABCA4,a gene that is too large to be packaged in AAV.Therefore,we investigated the ability of helper dependent adenovirus(HD-Ad)to deliver genes to the retina as it has a much larger transgene capacity.Using an EGFP reporter,our results showed that HD-Ad can transduce the entire retinal epithelium of a mouse using a dose of only 1
105 infectious units and maintain transgene expression for at least 4 months.The results demonstrate that HD-Ad has the potential to be an effective vector for the gene therapy of the retina.

Authentication of experimental materials:A remedy for the reproducibility crisis?

Reproducibility has always been a serious challenge when medical researchers in both academia and industry have tried to build upon previously published discoveries.Blindly chasing faulty results has incurred a huge waste of human and monetary resources.The damage to the progress of scientific discoveries,as well as their application to human well-being,cannot be overestimated.According to two reports by Bayer and Amgen published in 2011 and 2012,64e89%of the socalled“landmark”results could not be reproduced in their pre-clinical validation experiments.1,2 One plausible explanation for this out of proportion irreproducibility is related to the intricacy of the scientific experiments,including the sourcing of reagent antibodies and cell lines,which are major sources of variations.To make validation meaningful,the study materials used in the original studies need to be authenticated so that variations due to the faulty materials can be prevented during follow-up studies.However,the technical complexity and the costs of authentication often discourage this practice in research laboratories.

Seeing is believing:Stem cells to treat blindness

The majority of clinical blindness is caused by a loss of transparency of the lens and cornea,largely due to cataracts and corneal injuries.The most common treatment used to restore the transparency is surgical removal of the damaged tissues,followed by transplantation of donated corneal tissue or an artificial lens.However,these therapies are not without limitations or untoward effects.Unraveling the intricate regulatory signals required for cornea and lens development has made it possible to harness the lineage growth potential of stem cells for cornea repair and lens regeneration,as showcased in two recent studies published in the March 17th issue of Nature.

iPSC-based treatment of age-related macular degeneration (AMD):The path to success requires more than blind faith

Induced pluripotent stem cells(iPSCs)hold great promise for the treatment of human diseases.Two recent first-of-its-kind clinical case reports on the iPSC-based treatment of age-related macular degeneration(AMD)highlight the hopes and challenges associated with the clinical application of iPSCs.

Gene editing:A new step and a new direction toward finding a cure for Duchenne muscular dystrophy(DMD)

Duchenne muscular dystrophy(DMD)is a progressive muscle degenerative disease affecting one out of 3500 male births.Patients usually succumb to the disease by age 25.It has been shown that skipping exons of the DMD gene that contain disease-causing mutations from the pre-mRNA can result in a shortened,but functional,dystrophin protein that could bring clinical benefits to patients.A recent breakthrough has been reported in Science by three groups who demonstrated that genetically deleting exon 23 by gene editing can restore the expression of dystrophin(albeit a shortened version)and improve the muscle function in a mouse model of DMD.

CRISPR comes of age:A fairytale turned into bedside reality?

We devote this short piece to highlight one recent article published in Cell Stem Cell,reporting the correction of large chromosomal inversions of the factor VIII(F8)gene in cells from Hemophilia A patients using the CRISPR-Cas9 technology,one of the first attempts to edit large segments of chromosomes in patient cells using such methodology.The corrected cells were found free of off-target mutations and producing functional factor VIII in hemophilia mouse model.This work heralds another major advance in bringing CRISPR closer to the therapeutic reality.

Direct lineage conversion with pluripotency factors:A risky detour through transient pluripotency?

The advent of induced pluripotent stem cells(iPSCs)marked a giant step forward towards the reality of converting one type of primary somatic cells into different lineages capable of clinically repairing damaged tissues and organs.However,the major drawbacks of iPSCs hinder their quick translation to the bedside.These drawbacks include the time-,cost-,and labor-intensive process in production of clinical products from iPSCs,and the inherent risk of long-term tumorigenesis due to the forced expression of transcription factors associated with pluripotency,which are often implicated as aberrations within the cancerous gene circuitry.