The combination of forskolin and VPA increases gene expression efficiency to the hypoxia/neuron-specific system

ABSTRACT PHAX (phosphorylated adaptor for RNA export) is the key regulator of U snRNA nuclear export in metazoa. Our previous work revealed that PHAX is phosphorylated in the nucleus and is exported as a component of the U snRNA export complex to the cytoplasm, where it is dephosphorylated (M. Ohno, A. Segref, A. Bachi, M. Wilm, and I. W. Mattaj, Cell 101:187-198, 2000). PHAX phosphorylation is essential for export complex assembly, whereas its dephosphorylation causes export complex disassembly. Thus, PHAX is subject to a compartmentalized phosphorylation/dephosphorylation cycle that contributes to transport directionality. However, neither essential PHAX phosphorylation sites nor the modifying enzymes that contribute to the compartmentalized system have been identified. Here, we identify PHAX phosphorylation sites that are necessary and sufficient for U snRNA export. Mutation of the phosphorylation sites inhibited U snRNA export in a dominant-negative way. We also show, by both biochemical and RNA interference knockdown experiments, that the nuclear kinase and the cytoplasmic phosphatase for PHAX are CK2 kinase and protein phosphatase 2A, respectively. Our results reveal the composition of the compartmentalized phosphorylation/dephosphorylation system that regulates U snRNA export. This finding was surprising in that such a specific system for U snRNA export regulation is composed of two such universal regulators, suggesting that this compartmentalized system is used more broadly for gene expression regulation.

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An Evaluation on Transfection Efficiency of pHRE-Egr1-EGFP in Hepatocellular Carcinoma Cells Bel-7402 Mediated by PEI-MZF-NPs

Journal of Nanomaterials ◽

10.1155/2011/136052 ◽

2011 ◽

Vol 2011 ◽

pp. 1-10 ◽

Cited By ~ 2

Author(s):

Mei Lin ◽

Dongsheng Zhang ◽

Junxing Huang ◽

Jia Zhang ◽

Li Wang ◽

...

Keyword(s):

Gene Expression ◽

Hepatocellular Carcinoma ◽

Target Gene ◽

Transfection Efficiency ◽

Egfp Expression ◽

Egfp Gene ◽

Expression Efficiency ◽

Induction Components ◽

Eukaryotic Gene ◽

Viable Approach

To improve transfection and expression efficiency of target gene, especially under cancer anoxic microenvironment, we have developed pHRE-Egr1-EGFP/PEI-MZF-NPs nanosystem, in which pHRE-Egr1-EGFP, eukaryotic gene expression plasmid, is constructed by combining radiation promoter Egr1 with anoxia induction components (HRE), forming anoxic radiation double sensitive HRE/Egr1 promoter to activate reporter gene EGFP expression. MZF-NPs (Mn0.5Zn0.5Fe2O4magnetic nanoparticles), obtained by coprecipitation method, are coated with cation poly(ethylenimine) (PEI). We transferred pHRE-Egr1-EGFP into hepatocellular carcinoma Bel-7402 cells, using PEI-MZF-NPs as the carrier and tested some relevant efficacy. The results show that PEI-MZF-NPs have good DNA-binding ability, protection ability, release ability, little toxicity, and high transfection efficiency, obviously superior to those of the liposome method and electricity perforation method. Moreover, the expression level of EGFP gene induced by anoxia and radiation was significantly higher than that of single radiation activation. It is therefore concluded that HRE/Egr1 can induce and improve target gene expression efficiency in cancer anoxic microenvironment, and that PEI-MZF-NPs can be used as a novel nonviral gene vector which offers a viable approach to the mediated radiation gene therapy of cancer.

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A HIGHLY SENSITIVE AND SPECIFIC SYSTEM FOR LARGE-SCALE GENE EXPRESSION PROFILING.

Journal of Investigative Medicine ◽

10.1097/00042871-200703010-00052 ◽

2007 ◽

Vol 55 (2) ◽

pp. S356

Author(s):

G. Hu ◽

Q. Yang ◽

H. Li

Keyword(s):

Gene Expression ◽

Gene Expression Profiling ◽

Expression Profiling ◽

Large Scale ◽

Specific System ◽

Highly Sensitive

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Comparison of gene expression efficiency and innate immune response induced by Ad vector and lipoplex

Journal of Controlled Release ◽

10.1016/j.jconrel.2006.11.030 ◽

2007 ◽

Vol 117 (3) ◽

pp. 430-437 ◽

Cited By ~ 31

Author(s):

Haruna Sakurai ◽

Fuminori Sakurai ◽

Kenji Kawabata ◽

Tomomi Sasaki ◽

Naoya Koizumi ◽

...

Keyword(s):

Gene Expression ◽

Immune Response ◽

Innate Immune Response ◽

Innate Immune ◽

Expression Efficiency

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Heterologous Gene Expression in a Membrane-Protein-Specific System

Protein Expression and Purification ◽

10.1006/prep.1999.1110 ◽

1999 ◽

Vol 17 (2) ◽

pp. 312-323 ◽

Cited By ~ 22

Author(s):

George J. Turner ◽

Regina Reusch ◽

Ann M. Winter-Vann ◽

Lynell Martinez ◽

Mary C. Betlach

Keyword(s):

Gene Expression ◽

Membrane Protein ◽

Heterologous Gene Expression ◽

Heterologous Gene ◽

Specific System

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Physical stability and in-vitro gene expression efficiency of nebulised lipid–peptide–DNA complexes

International Journal of Pharmaceutics ◽

10.1016/s0378-5173(00)00339-2 ◽

2000 ◽

Vol 197 (1-2) ◽

pp. 221-231 ◽

Cited By ~ 49

Author(s):

James C Birchall ◽

Ian W Kellaway ◽

Mark Gumbleton

Keyword(s):

Gene Expression ◽

Physical Stability ◽

Dna Complexes ◽

Expression Efficiency

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A highly sensitive and specific system for large-scale gene expression profiling

BMC Genomics ◽

10.1186/1471-2164-9-9 ◽

2008 ◽

Vol 9 (1) ◽

pp. 9 ◽

Cited By ~ 3

Author(s):

Guohong Hu ◽

Qifeng Yang ◽

Xiangfeng Cui ◽

Gang Yue ◽

Marco A Azaro ◽

...

Keyword(s):

Gene Expression ◽

Gene Expression Profiling ◽

Expression Profiling ◽

Large Scale ◽

Specific System ◽

Highly Sensitive

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Self-organization controls expression more than abundance of molecular components of transcription and translation in confined cell-free gene expression

10.1101/401794 ◽

2018 ◽

Cited By ~ 1

Author(s):

P.M. Caveney ◽

R. Dabbs ◽

G. Chauhan ◽

S.E. Norred ◽

C.P. Collier ◽

...

Keyword(s):

Gene Expression ◽

Protein Production ◽

Self Organization ◽

Large Variability ◽

Expression Noise ◽

Expression Efficiency ◽

Cell Extracts ◽

Reaction Chambers ◽

Free Gene ◽

Molecular Components

AbstractCell-free gene expression using purified components or cell extracts has become an important platform for synthetic biology that is finding a growing numBer of practical applications. Unfortunately, at cell-relevant reactor volumes, cell-free expression suffers from excessive variability (noise) such that protein concentrations may vary by more than an order of magnitude across a population of identically constructed reaction chambers. Consensus opinion holds that variability in expression is due to the stochastic distribution of expression resources (DNA, RNAP, ribosomes, etc.) across the population of reaction chambers. In contrast, here we find that chamber-to-chamber variation in the expression efficiency generates the large variability in protein production. Through analysis and modeling, we show that chambers self-organize into expression centers that control expression efficiency. Chambers that organize into many centers, each having relatively few expression resources, exhibit high expression efficiency. Conversely, chambers that organize into just a few centers where each center has an abundance of resources, exhibit low expression efficiency. A particularly surprising finding is that diluting expression resources reduces the chamber-to-chamber variation in protein production. Chambers with dilute pools of expression resources exhibit higher expression efficiency and lower expression noise than those with more concentrated expression resources. In addition to demonstrating the means to tune expression noise, these results demonstrate that in cell-free systems, self-organization may exert even more influence over expression than the abundance of the molecular components of transcription and translation. These observations in cell-free platform may elucidate how self-organized, membrane-less structures emerge and function in cells.

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