Gene transfer technology is being used to enhance agronomic performance or improve quality traits in a wide variety of crop species. However, it is sometimes severely handicapped by difficulty in obtaining material in which transgene expression is predictable and stable over many generations. Because integration seemed to occur randomly in the plant genome, it was thought that some transgenes would be integrated in a relatively uncondensed, transcriptionally active chromatin environment, while others in a condensed, transcriptionally inert chromatin structure. Nuclear matrix attachment regions (MARs) are defined as DNA sequences that bind preferentially to the proteins of the nuclear matrix. They typically are localized at the borders of gene domains, implicating them in the formation of individual loops of higher order chromatin structure and transcription regulation. When MARs are positioned on either side of a transgene their presence usually results in higher and more stable espression in transgenic plants, most likely by minimizing gene silencing. In this review, we focus mainly on novel findings and our observations concerning the function of MARs in transcription regulation. Our objective is not only to summarize the current data and present several possible models to explain MAR effects on the transcription regulation, but also to point out some open questions involving the utilization of MARs in constructing high efficient expression vectors.
Chromatin ; physiology ; DNA ; metabolism ; Gene Expression Regulation, Plant ; Models, Genetic ; Nuclear Matrix ; metabolism ; Nuclear Proteins ; metabolism ; Transcription, Genetic ; Transgenes ; genetics

Peroxisome proliferation is a cellular response to many chemical compounds affects including natural and modified fatty acids, phthalate and adipate ester plasticizers, leukotriene antagonists, acetylsalicylic acid and certain pathophysiological conditions including dramatic change of cellular morphology and enzymatic activity. Peroxisome proliferation phenomenon is seen primarily in liver and kidney. Hormones and nutritional factor can regulate peroxisome proliferation response. Sustained peroxisome proliferation can lead to hepatocarcinogenesis. The three types of peroxisome proliferator activated receptor, termed PPAR alpha, PPAR beta, and PPAR gamma, expressed in specific tissue, are consisted of a specific a nuclear receptor superfamily. After more than 10 years world wide research, the function of PPAR is clarified, as PPAR gamma, the master of thrifty genes, controls the expression of genes relative to adipogenesis, diabetes mellitus and obesity. The receptor is involved in transcriptional control of numerous cellular processes including cell cycle control, inflammation, immunoregulation and carcinogenesis.
Adipocytes ; cytology ; Animals ; Cell Differentiation ; Energy Metabolism ; genetics ; Humans ; Intracellular Signaling Peptides and Proteins ; Nuclear Receptor Coactivators ; Peroxisome Proliferators ; Receptors, Cytoplasmic and Nuclear ; genetics ; physiology ; Transcription Factors ; genetics ; physiology

Enterovirus 71 (EV71) is a neurotropic pathogen that can induce hand, foot and mouth disease in children. There is an appreciable mortality rate after EV71 infections. The mechanism of action of EV71 replication is not known. Recent work has identified some of cell factors of the host that participate in the synthesis of the RNA and proteins of EV71 (e.g., hnRNP K, reticulon 3 (RTN 3)). In that work, researchers used a competitive assay to show that hnRNP K can interact with EV71 5' UTR, which is required for efficient synthesis of viral RNA. Using a yeast two-hybrid system, other researchers demonstrated that RTN 3 interacts with the N-terminal domain of EV71 2C, which is crucial for replication of viral RNA. Here, we discuss recent work focusing on the molecular mechanisms of hnRNP K and RTN 3 in the synthesis of the RNA and proteins of EV71.
Animals ; Carrier Proteins ; genetics ; metabolism ; Enterovirus A, Human ; genetics ; physiology ; Enterovirus Infections ; genetics ; metabolism ; virology ; Heterogeneous-Nuclear Ribonucleoprotein K ; Host-Pathogen Interactions ; Humans ; Membrane Proteins ; genetics ; metabolism ; Nerve Tissue Proteins ; genetics ; metabolism ; Ribonucleoproteins ; genetics ; metabolism ; Viral Proteins ; genetics ; metabolism ; Virus Replication

Animals ; Apoptosis ; Cell Cycle ; Cell Cycle Proteins ; genetics ; metabolism ; physiology ; DNA Repair ; Homeodomain Proteins ; genetics ; metabolism ; physiology ; Humans ; Inhibitor of Growth Protein 1 ; Intracellular Signaling Peptides and Proteins ; genetics ; metabolism ; physiology ; Neoplasm Metastasis ; Neoplasms ; metabolism ; pathology ; Neovascularization, Pathologic ; pathology ; Nuclear Proteins ; genetics ; metabolism ; physiology ; Prognosis ; Receptors, Cytoplasmic and Nuclear ; genetics ; metabolism ; physiology ; Signal Transduction ; Transcription Factors ; genetics ; metabolism ; physiology ; Tumor Suppressor Protein p53 ; metabolism ; Tumor Suppressor Proteins ; chemistry ; genetics ; metabolism ; physiology

We investigated the microRNA172 (miR172)-mediated regulatory network for the perception of changes in external and endogenous signals to identify a universally applicable floral regulation system in ornamental plants, manipulation of which could be economically beneficial. Transgenic gloxinia plants, in which miR172 was either overexpressed or suppressed, were generated using Agrobacterium-mediated transformation. They were used to study the effect of altering the expression of this miRNA on time of flowering and to identify its mRNA target. Early or late flowering was observed in transgenic plants in which miR172 was overexpressed or suppressed, respectively. A full-length complementary DNA (cDNA) of gloxinia (Sinningia speciosa) APETALA2-like (SsAP2-like) was identified as a target of miR172. The altered expression levels of miR172 caused up- or down-regulation of SsAP2-like during flower development, which affected the time of flowering. Quantitative real-time reverse transcription PCR analysis of different gloxinia tissues revealed that the accumulation of SsAP2-like was negatively correlated with the expression of miR172a, whereas the expression pattern of miR172a was negatively correlated with that of miR156a. Our results suggest that transgenic manipulation of miR172 could be used as a universal strategy for regulating time of flowering in ornamental plants.
Arabidopsis/genetics* ; Arabidopsis Proteins/metabolism* ; Cloning, Molecular ; DNA, Complementary/metabolism* ; Flowers/physiology* ; Gene Expression Profiling ; Gene Expression Regulation, Plant ; Homeodomain Proteins/metabolism* ; Lamiales/physiology* ; MicroRNAs/metabolism* ; Nuclear Proteins/metabolism* ; Plants, Genetically Modified/physiology* ; Plasmids/metabolism* ; Polymerase Chain Reaction ; Transgenes

AIMTo investigate the roles of liver X receptors (LXR) in the lipid composition and gene expression regulation in the murine caput epididymidis. LXR are nuclear receptors for oxysterols, molecules derived from cholesterol metabolism that are present in mammals as two isoforms: LXRalpha, which is more specifically expressed in lipid-metabolising tissues, such as liver, adipose and steroidogenic tissues, and macrophages, whereas LXRbeta is ubiquitous. Their importance in reproductive physiology has been sustained by the fact that male mice in which the function of both LXR has been disrupted have fertility disturbances starting at the age of 5 months, leading to complete sterility by the age of 9 months. These defects are associated with epididymal epithelial degeneration in caput segments one and two, and with a sperm midpiece fragility, leading to the presence of isolated sperm heads and flagella when luminal contents are recovered from the cauda epididymidis.

METHODSThe lipid composition of the caput epididymidis of wild-type and LXR-deficient mice was assessed using oil red O staining on tissue cryosections and lipid extraction followed by high performance liquid chromatography or gas chromatography. Gene expression was checked by quantitative real time polymerase chain reaction.

RESULTSUsing LXR-deficient mice, we showed an alteration of the lipid composition of the caput epididymidis as well as a significantly decreased expression of the genes encoding SREBP1c, SCD1 and SCD2, involved in fatty acid metabolism.

CONCLUSIONAltogether, these results show that LXR are important regulators of epididymal function, and play a critical role in the lipid maturation processes occurring during sperm epididymal maturation.

Animals ; DNA Primers ; DNA-Binding Proteins ; deficiency ; genetics ; physiology ; Epididymis ; cytology ; physiology ; Epithelial Cells ; physiology ; Fatty Acids ; metabolism ; Homeostasis ; Lipids ; physiology ; Liver X Receptors ; Male ; Mice ; Mice, Knockout ; Orphan Nuclear Receptors ; Polymerase Chain Reaction ; Receptors, Cytoplasmic and Nuclear ; deficiency ; genetics ; physiology

OBJECTIVETo explore the effect of a non-lethal heat shock, in comparison with the treatment of interferon-gamma (IFN gamma), on the expression of major histocompatibility complex transactivator (CTA) and its downstream target gene of the human leukocyte antigens (HLA)-DR in Jurkat cells.

METHODSThe changes of CTA mRNA in Jurkat cells before and after the treatment of heat shock or IFN gamma were detected using real time RT-PCR. The changes of CTA protein were detected with Western blot. The expression of HLA-DR was detected with flow cytometry. : CTA mRNA and protein were induced in Jurkat cells under heat shock, but not with IFN-gamma. The expression of HLA-DR gene significantly increased after recovery (P<0.01).

CONCLUSIONThe expressions of CTA and HLA-DR in Jurkat cells remarkably increase after heat shock, indicating that heat shock may help reconstruct relevant genes in cells with immunologic gene deficiencies.

HLA-DR Antigens ; metabolism ; Heat-Shock Response ; physiology ; Humans ; Jurkat Cells ; Nuclear Proteins ; genetics ; metabolism ; RNA, Messenger ; genetics ; Trans-Activators ; genetics ; metabolism

OBJECTIVETo investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells.

METHODSThe binding of trans-acting factor(s) to two enhancers of the rGSTP1 gene, glutathione S-transferase P enhancer I (GPEI) and glutathione S-transferase P enhancer II-1 (GPE II-1), was identified by an electrophoretic mobility shift assay (EMSA). The molecular weight of trans-acting factor was measured in a UV cross-linking experiment.

RESULTSTrans-acting factor interacting with the core sequence of GPEI (cGPEI) were found in human cervical adenocarcinoma cell line (HeLa) and rat hepatoma cell line (CBRH7919). These proteins were not expressed in normal rat liver. Although specific binding proteins that bound to GPE II-1 were detected in all three cell types, a 64 kDa binding protein that exists in HeLa and CBRH7919 cells was absent in normal rat liver.

CONCLUSIONcGPEI, GPEII specific binding proteins expressed in HeLa and CBRH7919 cells may play an important role in the high transcriptional level of the rGSTP1 gene in tumor cells.

Animals ; Carrier Proteins ; metabolism ; Enhancer Elements, Genetic ; physiology ; Gene Expression Regulation, Enzymologic ; Glutathione S-Transferase pi ; Glutathione Transferase ; genetics ; Isoenzymes ; genetics ; Nuclear Proteins ; metabolism ; Rats ; Transcription, Genetic

Bel1, a transactivator of prototype foamy virus (PFV), plays pivotal roles in the replication of PFV. Previous studies have shown that Bel1 bears a nuclear localization signal (NLS), but its amino acid sequence remains unclear and the corresponding importins have not been identified. In this report, we inserted various fragments of Bel1 into an EGFP-GST fusion protein and investigated their subcellular localization by fluorescence microscopy. We found that the 215PRQKRPR221 fragment could direct nuclear localization, which accords with the consensus sequence K(K/R)X(K/R) of monopartite NLS. Point mutation experiments revealed that K218, R219, and R221 are essential for the nuclear localization of Bel1. The results of the GST-pulldown showed that the Bel1 fragment with residues 215-223, which bears the NLS, interacts with KPNA1, KPNA6, and KPNA7. This result suggests that KPNA1, KPNA6, and KPNA7 maybe involved in Bel1 nuclear translocation.
Cell Line ; Cell Nucleus ; genetics ; metabolism ; virology ; Humans ; Nuclear Localization Signals ; genetics ; metabolism ; Protein Binding ; Protein Transport ; Retroviridae Infections ; genetics ; metabolism ; virology ; Retroviridae Proteins ; chemistry ; genetics ; metabolism ; Spumavirus ; chemistry ; genetics ; physiology ; Trans-Activators ; chemistry ; genetics ; metabolism ; alpha Karyopherins ; genetics ; metabolism

Sequential activation of the JNK pathway components, including Rac1/Cdc42, MLKs (mixed-lineage kinases), MKK4/7 and JNKs, plays a required role in many cell death paradigms. Those components are organized by a scaffold protein, POSH (Plenty of SH3's), to ensure the effective activation of the JNK pathway and cell death upon apoptotic stimuli. We have shown recently that the expression of POSH and MLK family proteins are regulated through protein stability. By generating a variety of mutants, we provide evidence here that the Nterminal half of POSH is accountable for its stability regulation and its over-expression-induced cell death. In addition, POSH's ability to induce apoptosis is correlated with its stability as well as its MLK binding ability. MLK family's stability, like that of POSH, requires activation of JNKs. However, we were surprised to find out that the widely used dominant negative (d/n) form of c-Jun could down-regulate MLK's stability, indicating that peptide from d/n c-Jun can be potentially developed into a therapeutical drug.
Adaptor Proteins, Signal Transducing ; genetics ; metabolism ; Animals ; Apoptosis ; physiology ; Base Sequence ; Cell Line ; DNA Primers ; genetics ; Humans ; JNK Mitogen-Activated Protein Kinases ; antagonists & inhibitors ; genetics ; metabolism ; MAP Kinase Kinase Kinases ; genetics ; metabolism ; Mutant Proteins ; genetics ; metabolism ; Nuclear Proteins ; genetics ; metabolism ; PC12 Cells ; Peptide Fragments ; genetics ; metabolism ; Protein Stability ; Rats ; Recombinant Proteins ; genetics ; metabolism ; Signal Transduction ; physiology ; Transfection ; Ubiquitin-Protein Ligases ; genetics ; metabolism