Effect on Survivin Regulation of Transcription Level by p21^(waf1) Overexpression in HepG2 Hepatocellular Carcinoma Cells

The effect of cyclin-dependent kinase inhibitors Cip1/Wafl （p21） on regulatory expression of survivin transcription in human hepatocellular carcinoma cell HepG2 was observed and the related mechanisms explored. Doxorubicin （DOX） was used to treat HepG2. Eukaryotic vector pEGFP-C2-p21 was transfected into HepG2 by lipofectamine and positive clones were screened out by G418. The mRNA expression of p21 and survivin was detected by real-time fluorescent quantitative polymerase chain reaction （RQ-PCR）. Flow cytometry was used to examine the cell cycle, and reverse transcription polymerase chain reaction （RT-PCR） was used to measure the levels of E2F-1 and p300. The results showed that： （1） After treatment with DOX, the expression of p21 was increased, whereas that of survivin was reduced during 24 h of treatment; （2） After transfection of pEGFP-C2-p21 into HepG2, p21 level was significantly enhanced to 2100.11-folds or 980.89-folds in comparison to HepG2 or HepG2-C2 group, and survivin level was markedly down-regulated to 0.54% or 0.59% relative to the control groups; （3） Overexpressed p21 resulted in GI/G0 phase arrest （F=31.59, P〈0.01）, meanwhile E2F-1 mRNA and p300 mRNA were reduced as compared with those of controls （FE2F-1=I25.28, P〈0.05; Fp300= 46.01, P〈0.01）. It was suggested that p21 could be a potential mediator of survivin suppression at transcription level in HepG2 cell, which might be through the block at G1/G0 phase and down-regulation of transcription factors E2F-1 and p300.

A Study on the Molecular Switch of Gene Expression of the Mouse Heart Nuclear DNA Fragments

It is observed by in situ stain that LDH (1 5) ...nNAD + can probably enter the nucleopore and can be bound bound specifically with the genes that encode them. During the in vitro expression, the dilution of heart nuclear DNA fragments could enhance the expression activity of LDH/DNA and the amount of expressed LDH (1 5) is in proportion to the amount of dissociable LDH (1 5) on the LDH/DNA. With the integration of 14C Leu to the proteins, it is also observed that the addition of LDH (1 5) ...nNAD + can suppress the in vitro expression activity of LDH/DNA. AFM observation shows that the regulation sequence at the both ends of active genes may be bound with such active factors as proteins encoded by the genes which probably is the main molecular switch of gene expression and regulation we have been always searching for. Our work shows the prospective application of the combination of AFM and isotope labeling in the research of biological reaction.

Responses Taken by Silencing of NFkappaB, STAMP1 and STAMP2 Genes and Expression of NFkB, Act-1, p53 and p73 at -/+ TNFalpha Induced LNCaP Cells

Prostate cancer is the most commonly diagnosed cancer and the second leading cause of cancer mortality in men in the Western World. The effects of androgens are mediated by the Androgen Receptor (AR). Therefore, studies focus on the identification of AR-regulated genes that are also highly expressed in the prostate. STAMP family genes STAMP1/STEAP2 and STAMP2/STEAP4 have only expressed in androgen receptor-positive cells, the role of AR in STAMP family gene expression is an important question. STEAP (Six Transmembrane Epithelial Antigens of Prostate) is the first characterized prostate enriched six transmembrane genes, expressed in metastatic prostate cancer samples, it is tempting to speculate that STAMP/STEAP family genes may be involved in similar functions with a role for both the normal biology and pathophysiology of the prostate. Using siRNA technology in LNCaP cells expressing STAMP genes per se, an apoptosis panel including pro-apoptotic and/or apoptotic molecules was assayed by RT-PCR. In this research project, the prostate-specific STAMP gene family and its regulatory effects on the nuclear factor kappa B and caspase-related pathways were characterized. Considering that the beta-actin response in the control group was high in the immunolabeling studies, an increase in the induction of Tumor Necrosis Factor (TNF) was detected in the signals received with the vital proteins NFkB and akt, which were silenced by siRNA, which means that STAMP genes potentiate vital proteins.

Regulatory role of NFAT1 signaling in articular chondrocyteactivities and osteoarthritis pathogenesis

Osteoarthritis (OA), the most common form of joint disease, is characterized clinically by joint pain, stiffness,and deformity. OA is now considered a whole joint disease;however, the breakdown of the articular cartilage remains themajor hallmark of the disease. Current treatments targeting OA symptoms have a limited impact on impeding orreversing the OA progression. Understanding the molecular and cellular mechanisms underlying OA development isa critical barrier to progress in OA therapy. Recent studies by the current authors’ group and others have revealedthat the nuclear factor of activated T cell 1 (NFAT1), a member of the NFAT family of transcription factors, regulatesthe expression of many anabolic and catabolic genes in articular chondrocytes of adult mice. Mice lacking NFAT1exhibit normal skeletal development but display OA in both appendicular and spinal facet joints as adults. Thisreview mainly focuses on the recent advances in the regulatory role of NFAT1 transcription factor in the activities ofarticular chondrocytes and its implication in the pathogenesis of OA.

Effects of Oxidized Low Density Lipoprotein onthe Expression and Function of ABCA1 in Macrophages

Summary:In the present study, we examined the regulation of the expression and function of ABCA1 by modified LDL (ox-LDL) in vitro. After incubation with apoA-I for 24 h, RAW264.7 cells effluxed 37.65 % cholesterol loaded by acetyl LDL (ac-LDL), and 9.78 % cholesterol in ox-LDL group. The level of ABCA1 mRNA increased about three times either when cells were incubated with 100 μg /mL ac-LDL or with 100 μg /mL ox-LDL. However, the level of ABCA1 protein rose by 1.57 times in ac-LDL group and 1.26 times in ox-LDL group. These results demonstrated that ox-LDL had different effect on the expression and function of ABCA1, ox-LDL might decrease the cholesterol efflux mediated by ABCA1 through other unknown mechanisms.

EBNA3C combines with Gemin3 and up-regulates its expression

Objective To investigate the combination of Epstein-Barr virus nuclear protein 3C (EBNA3C) with Gemin3 and its effect on Gemin3 expression. Methods Co-immunoprecipitation, GST pull-down and immunofluorescent assay were used to determine the combination of EBNA3C and Gemin3 and their combining domain. Stable EBNA3C knockdown cell lines were made by lentivirus-delivered small hairpin RNA and then puromycin selection. Western blot was used to check the effect of EBNA3C on Gemin3 expression. Results EBNA3C and Gemin3 combined with each other in vivo and in vitro through their C-terminals. EBNA3C up-regulated Gemin3 gene expression. Conclusion EBNA3C forms complex with Gemin3 and up-regulates its expression.

MicroRNAs Involved in the Pathogenesis of Phytophthora Root Rot of Soybean (Glycine max)

Phytophthora root rot is one of the most prevalent diseases in the world,which can infect the seedlings and plants,with substantial negative impact on soybean yield and quality.MicroRNAs （miRNAs） are a class of post-transcriptional regulators of gene expression during growth and development of organisms.A soybean disease-resistance variety Suinong 10 was inoculated with Phytophthora sojae race No.1,and the specific miRNA resistant expression profile was acquired by microarray for the first time.Different expressional miRNAs have been found after comparing the results of the treated sample with the control sample.Furthermore,the target genes of different expressional miRNAs were predicted.Two miRNAs,cbr-mir-241 and ath-miR854a,regulated the disease-resistance process directly through their targets,some enzymes.Another two miRNAs,gma-miR169a and ath-miR169h,participated in disease-resistance regulation as transcription factors.Similarly,one miRNA,ptc-miR164f,has been reported to regulate the plant development.All of these studies would be served as the foundation for exploring the resistance mechanism.

Flow-dependent regulation of vascular function and gene expression in rat superior mesenteric artery

Background Mesenteric artery thrombosis is prone to occur at specific arterial regions with different fluid flow patterns, yet mechanistic links between blood flow and vascular function remain unclear. This study aimd to investigate the role of blood flow in regulation of vascular function and gene expression in rats. Methods isometric tension was recorded in wire myograph to examine vascular function of specific regions （trunk parts and proximal parts from the origin） with different blood flow in superior mesenterJc artery （SMA）. Endothelial nitric oxide syntheses （eNOS）, phosphorylated-eNOS （p-eNOS）, serine-threonine kinase Akt and phosphorylated-Akt （p-Akt） protein expressions in SMA were examined by Western blotting. Significance was analyzed using a Student＇s t test or analysis of variance （ANOVA） followed by a Dunnett＇s multiple-comparison post hoc test. Results Compared with trunks, proximal parts exhibited severely impaired relaxant responses to acetylcholine （Ach） （1 nmol/L to 10 μmol/L） （P 〈0.01）. p-eNOS and p-Akt protein levels were significantly reduced in proximal parts of SMA （0.37±0.03, 0.42±0.03 respectively） versus trunk parts （0.82±0.03, 0.72±0.03 respectively, both P 〈0.05） while total eNOS and Akt expressions remain comparable in both regions by Western blotting analysis （0.70±0.03 vs 0.82±0.03; 0.70±0.03 vs 0.77±0.03 respectively, both P 〉0.05）. Conclusion Critical components that drive the vascular function and influence the localization of mesenteric artery thrombosis are flow-responsive elements within the vascular endothelium.

The role of microRNA-200a in the occurrence and development of liver disease

microRNA(miRNA)is a type of small non-coding RNA that can participate in cell proliferation and apoptosis by regulating gene expression.More and more evidences indicate that miRNA-200a is involved in the occurrence and development of non-alcoholic fatty liver disease,alcoholic liver disease,drug-induced liver injury,liver fibrosis,and hepatocellular carcinoma.Downstream target genes of serotonin,regulating related signal pathways and playing different roles in the progression of a variety of liver diseases,provide a reference for exploring the mechanism of a variety of chronic liver diseases.

Involvement of transcriptional enhancers in the regulation of developmental expression of yellow gene

Upstream regulatory region and flanking DNA of yellow gene wereisolated and cloned from a Drosophila genomic library. A vector containing yellow gene and regulatory elements was constructed using the recombinant DNA technique. Then this vector was integrated into Drosophila genome by genetic transformation. Using both FLP/FRT and Cre/LoxP site-specific recombination systems, two new yellow alleles were created at the same position in the genome of transgenic flies. Results from genetic and molecular analysis indicated that transcriptional enhancers regulate the developmental expression of the transgene. Furthermore, interactions between new-created yellow alleles were observed. Such interactions can influence markedly the expression of yellow gene during development. This effect may also be a form of enhancer-mediated gene expression.

MeCP2:multifaceted roles in gene regulation and neural development

Methyl-CpG-binding protein 2 （MeCP2） is a classic methylated-DNA-binding protein, dysfunctions of which lead to various neurodevelopmental disorders such as Rett syndrome and autism spectrum disorder. Initially recognized as a transcriptional repressor, MeCP2 has been studied extensively and its functions have been expanded dramatically in the past two decades. Recently, it was found to be involved in gene regulation at the post-transcriptional level. MeCP2 represses nuclear microRNA processing by interacting directly with the Drosha/DGCR8 complex. In addition to its multifaceted functions, MeCP2 is remarkably modulated by post- translational modifications such as phosphorylation, SUMOylation, and acetylation, providing more regulatory dimensions to its functions. The role of MeCP2 in the central nervous system has been studied extensively, from neurons to glia. Future investigations combining molecular, cellular, and physiological methods are necessary for defining the roles of MeCP2 in the brain and developing efficient treatments for MeCP2-related brain disorders.

MSI/LOH and extron expression of the FHIT gene in gastric carcinoma

We detected loss of heterozygosity(LOH)and microsatellite instabilities(MSI),as well as extron expression of the fragile histidine triad(FHIT)gene in gastric carcinoma(GC),in order to evaluate their association with clinicopathological processes in gastric carcinogenesis.LOH and MSI of the FHIT were detected by using PCR at 4 microsatellite loci:D3S 1300,D3S 4103,D3S 1481,D3S 1234 in cancer tissues from 50 patients with primary GC,with normal mucosa acting as matched controls.FHIT transcripts were detected by nested RT-PCR in 30 cases of GC and their products were sequenced.Results show that the average frequencies of LOH and MSI of the FHIT gene in GC were 32.4%and 26.4%,respectively.There was no correlation between LOH and MSI of the FHIT gene in GC and the histological characteristics of gastric carcinoma(Bormann’s or Lauren’s classification).LOH of the FHIT gene in GC was related to depth invasiveness,and its frequency in GC where serosa was penetrated was significantly higher than that in GC without serosa penetration(73.5%vs 37.5%,P<0.05).The frequency of MSI in GC without lymph node metastasis was significantly higher than that in GC with lymph node metastasis(66.7%vs 34.3%,P<0.05).Aberrant transcripts were found in 11/30 GC tissues.Sequencing analysis of the aberrant fragments found a RT-PCR product missing exons 5–7 in one case of GC,and another product missing exons 4–7.Four of 10(40.0%)cases of primary GC showed absent or decreased expression of the FHIT protein as compared to their matched normal tissues.The findings in this study suggest that LOH and MSI of FHIT gene may induce aberrant extron expression,which might play a role in gastric carcinogenesis.

2型固有淋巴细胞调控因素的研究进展

2型固有淋巴细胞(group 2 innate lymphoid cells,ILC2s)是一群不表达谱系发育因子Lineage,不具有抗原特异性的特殊免疫细胞,能够在IL-33和IL-25的刺激下产生大量IL-5和IL-13,介导2型先天性免疫应答,在变应性和炎症性疾病中发挥重要作用。近年来ILC2s不同细胞亚型以及复杂的调控因子网络逐渐被发现和证实,这些研究结果拓展了对先天性免疫应答的认识,尤其是ILC2s参与变应原或病原体介导的气道炎症反应,为临床治疗提供了新的思路。

Transcriptional Regulation of the Ambient Temperature Response by H2A.Z Nucleosomes and HSF1 Transcription Factors in Arabidopsis

Temperature influences the distribution, range, and phenology of plants. The key transcriptional activators of heat shock response in eukaryotes, the heat shock factors （HSFs）, have undergone large-scale gene amplification in plants. While HSFs are central in heat stress responses, their role in the response to ambient temperature changes is less well understood. We show here that the warm ambient temperature transcriptome is dependent upon the HSFA1 clade ofArabidopsis HSFs, which cause a rapid and dynamic eviction of H2A.Z nucleosomes at target genes. A transcriptional cascade results in the activation of multiple downstream stress-responsive transcription factors, triggering large-scale changes to the transcriptome in response to elevated temperature. H2A.Z nucleosomes are enriched at temperature-responsive genes at non-inducible temperature, and thus likely confer inducibility of gene expression and higher responsive dynamics. We propose that the antagonistic effects of H2A.Z and HSF1 provide a mechanism to activate gene expression rapidly and precisely in response to temperature, while preventing leaky transcription in the absence of an activation signal.

SKA2/FAM33A:A novel gene implicated in cell cycle,tumorigenesis,and psychiatric disorders

SKA2(spindle and KT associated 2),also referred to as FAM33A(family with sequence similarity 33,member A),is a recently identified gene involved in cell cycle regulation,and growing evidence is implicating its roles in tumorigenesis and psychiatric disorders.It has been demonstrated that SKA2,along with its coworkers SKA1 and SKA3,constitutes the SKA complex which plays a critical role in the maintenance of the metaphase plate and/or spindle checkpoint silencing during mitosis.SKA2 is over-expressed both in cancer cell lines and clinical samples including small cell lung cancer and breast cancer,whereas downregulation of SKA2 is associated with depression and suicidal ideation.The expression of SKA2 is regulated by transcription factors including NF-kB and CREB,miRNAs as well as DNA methylation.In this review,we provide an overview of studies that reveal SKA2 gene and protein characteristics as well as physiological function,with a special focus on its transcription regulatory mechanisms,and also provide a summary regarding the translational opportunity of the SKA2 gene as a clinical biomarker for cancers and psychiatric disorders.

Comparative Analysis of Human Genes Frequently and Occasionally Regulated by m^6A Modification

The m^6A modification has been implicated as an important epitranscriptomic marker, which plays extensive roles in the regulation of transcript stability, splicing, translation, and localization. Nevertheless, only some genes are repeatedly modified across various conditions and the principle of m^6A regulation remains elusive. In this study, we performed a systems-level analysis of human genes frequently regulated by m^6A modification （m^6Afreq genes） and those occasionally regulated by m^6A modification （m^6Aocca genes）. Compared to the m^6Aocca genes, the m^6Afreq genes exhibit gene importance-related features, such as lower dN/dS ratio, higher protein-protein interaction network degree, and reduced tissue expression specificity. Signaling network analysis indicates that the m^6Afreq genes are associated with downstream components of signaling cascades, high-linked signaling adaptors, and specific network motifs like incoherent feed forward loops. Moreover, functional enrichment analysis indicates significant overlaps between the m^6Afreq genes and genes involved in various layers of gene expression, such as being the microRNA targets and the regulators of RNA processing. Therefore, our findings suggest the potential interplay between m^6A epitranscriptomic regulation and other gene expression regulatory machineries.

Coordinated regulation for nature products discovery and overproduction in Streptomyces

Streptomyces is an important treasure trove for natural products discovery.In recent years,many scientists focused on the genetic modification and metabolic regulation of Streptomyces to obtain diverse bioactive compounds with high yields.This review summarized the commonly used regulatory strategies for natural products discovery and overproduction in Streptomyces from three main aspects,including regulator-related strategies,promoter engineering,as well as other strategies employing transposons,signal factors,or feedback regulations.It is expected that the metabolic regulation network of Streptomyces will be elucidated more comprehensively to shed light on natural products research in the future.

Effects of hemin and thermal stress exposure on JWA expression

To investigate the expression of JWA after hemin and(or)thermal stress exposure,we treated K562(chronic myelogenous leukemia cells)cells with different doses of hemin and thermal stress using different exposure times.The expression of JWA protein was determined by Western blot analysis.Reverse transcription-polymerase chain reaction was carried out to determine JWA mRNA expression.JWA promoter transcription activity analysis was performed by chloramphenicol acetyl transferase-enzyme linked immunosorbent assay(CAT-ELISA).The expression of JWA protein was significantly increased by up to(3.23±0.57)folds compared to the control in K562 cells after hemin treatment(50μM for one week),and a similar pattern was observed in the cells after treatment with thermal stress(42℃)for 2 hours[increased by(8.00±1.73)folds].The expression of JWA mRNA was also significantly elevated by up to(1.37±0.06)folds in K562 cells treated with hemin(30μM for 48 hours),and a similar regulatory pattern[increased by(1.87±0.13)folds]was observed with thermal stress exposure(42℃)for 30 minutes.However,a combined antagonistic effect was observed in the treatment of K562 cells with hemin(30μM,48 h)followed by thermal stress(42℃,30 min).CAT-ELISA further confirmed that either hemin or thermal stress treatment could up-regulate JWA transcription activity,however,the effects could be counteracted partly by treatment with a combination of both.Hemin and thermal stress might regulate JWA expression via distinct intracellular signal transduction pathways.

Noncoding RNA-chromatin association:Functions and mechanisms

Pervasive transcription of the mammalian genome produces hundreds of thousands of noncoding RNAs(ncRNAs).Numerous studies have suggested that some of these ncRNAs regulate multiple cellular processes and play important roles in physiological and pathological processes.Notably,a large subset of ncRNAs is enriched on chromatin and participates in regulating gene expression and the dynamics of chromatin structure and status.In this review,we summarize recent advances in the functional study of chromatin-associated ncRNAs and mechanistic insights into how these ncRNAs associate with chromatin.We also discuss the potential future challenges which still need to be overcome in this field.

Functions of Circular RNAs in the Research of Reproductive and Developmental Medicine

Circular RNAs(circRNAs)represent a mysterious class of noncoding RNAs that are generated by the circularization of exons or introns and characterized as being highly stable and abundant.Although circRNAs have been studied for several decades,our knowledge of these molecules remains limited.With the development of innovative bioinformatic tools and sequencing methods,comprehensive studies of circRNAs are now available in the literature.There is emerging evidence to show that circRNAs play roles in the regulation of gene expression.In this review,several primary potential functions of circRNAs are summarized;these include binding to microRNA/RNA-binding proteins,inhibiting/promoting messenger RNA translation,and their own translation.