Regulation of Hepatitis C Virus Replication and Gene Expression by the MAPK-ERK Pathway

The mitogen activated protein kinases-extracellular signal regulated kinases （MAPK-ERK） pathway is involved in regulation of multiple cellular processes including the cell cycle. In the present study using a Huh7 cell line Conl with an HCV replicon, we have shown that the MAPK-ERK pathway plays a significant role in the modulation of HCV replication and protein expression and might influence IFN-a signalling. Epithelial growth factor （EGF） was able to stimulate ERK activation and decreased HCV RNA load while a MAPK-ERK pathway inhibitor U0126 led to an elevated HCV RNA load and higher NS5A protein amounts in Conl cells. It could be further demonstrated that the inhibition of the MAPK-ERK pathway facilitated the translation directed by the HCV internal ribosome entry site. Consistently, a U0126 treatment enhanced activity of the HCV reporter replicon in transient transfeetion assays. Thus, the MAPK-ERK pathway plays an important role in the regulation of HCV gene expression and replication. In addition, cyclin-dependent kinases （CDKs） downstream of ERK may also be involved in the modulation of HCV replication since roscovitine, an inhibitor of CDKs had a similar effect to that of U0126. Modulation of the cell cycle progression by cell cycle inhibitor or RNAi resulted consistently in changes of HCV RNA levels. Further, the replication of HCV replicon in Conl cells was inhibited by IFN-~z. The inhibitory effect of IFN-CZ could be partly reversed by pre-incubation of Con-1 cells with inhibitors of the MAPK-ERK pathway and CDKs. It could be shown that the MAPK-ERK inhibitors are able to partially modulate the expression of interferon-stimulated genes.

Progress in DNA Aptamers as Recognition Components for Protein Functional Regulation

Proteins play a central role in all domains of life,and precise regulation of their activity is essential for understanding the related biological processes and therapeutic functions.Nucleic acid aptamers,the molecular recognition components derived from systematic evolution of ligands by exponential enrichment(SELEX),can specifically identify proteins with antibody-like recognition characteristics and help to regulate their activity.This minireview covers the SELEX-based selection of protein-binding aptamers,membrane protein analytical techniques based on aptamer-mediated target recognition,aptamer-mediated functional regulation of proteins,including membrane receptors and non-membrane proteins(thrombin as a model),as well as the potential challenges and prospects regarding aptamer-mediated protein manipulation,aiming to supply some useful information for researchers in this field.

Effect of PRAK gene knockout on the proliferation of mouseembryonic fibroblasts

p38 regulated/activated protein kinase(PRAK)plays a key role in cell senescence and tumor suppression.The aim of this study was to investigate if PRAK had effect on cell proliferation.The growth of PRAK+/+and PRAK^(–/–)mouse embryonicfibroblast(MEF)cells was measured by methylthiazoletetrazolium(MTT)colorimetric assay,and the proportion of the cell number in different phases of the cell cycle was analyzed byflow cytometry.The growth curves showed that the growth rate was notably decreased,and cell double time was elongated in PRAK^(–/–)cells;moreover,the number of PRAK^(–/–)cells was decreased by 44.5%compared with that of PRAK+/+cells cultured for 96 h,suggesting that G2/M transition is inhibited in PRAK^(–/–)cells.Meanwhile,G1/S transition was also inhibited in PRAK^(–/–)cells,observed withflow cytometry analysis.The ratios of G0/G1,G2/M,and S phases of PRAK+/+cells were 44.9%,12.2%,and 42.9%,respec-tively,while those of PRAK^(–/–)cells were 55.3%,7.3%,and 37.4%,respectively.There were 23.1%increase and 12.7%decrease of the number of PRAK^(–/–)cells in G1 and S phases in comparison with that of PRAK+/+cells,respectively.Taken together,PRAK gene knockout in MEF cells leads to cell cycle arrest and proliferation inhibition.