RNA-induced silencing is a process which allows cells to regulate the synthesis of specific proteins. RNA silencing is promoted by the protein C3PO (component 3 of RISC). We have previously found that phospholipase Cß, which increases intracellular calcium levels in response to specific G protein signals, inhibits C3PO activity towards certain genes. Understanding the parameters that control C3PO activity and which genes are impacted by G protein activation would help predict which genes are more vulnerable to downregulation. Here, using a library of 1018 oligonucleotides, we show that C3PO binds oligonucleotides with structural specificity but little sequence specificity. Alternately, C3PO hydrolyzes oligonucleotides with a rate that is sensitive to substrate stability. Importantly, we find that oligonucleotides with higher Tm values are inhibited by bound PLCß. This finding is supported by microarray analysis in cells over-expressing PLCß1. Taken together, this study allows predictions of the genes whose post-transcriptional regulation is responsive to the G protein/phospholipase Cß/calcium signaling pathway.
Phospholipase C beta/metabolism , Promoter Regions, Genetic/genetics , RNA-Induced Silencing Complex/metabolism , Gene Expression Regulation , Humans , Nucleic Acid Conformation , Oligonucleotide Array Sequence Analysis , Oligonucleotides/chemistry , Oligonucleotides/metabolism , RNA-Induced Silencing Complex/chemistry , RNA-Induced Silencing Complex/genetics
The predictive capabilities of models that satisfy the Weiner bounds and Hashin-Shtrikman (HS) bounds were studied for isotropic dielectrics in a laminar heterogeneous configuration oriented perpendicular to the electric field. The dynamics were investigated isothermally using broadband dielectric relaxation spectroscopy (DRS) in the frequency range of 1 MHz to 100 mHz. The molecules chosen for study were low molecular weight glass formers, glycerol, phenyl salicylate, imidazole, and dimethyl sulfoxide, and macromolecules, polymethylhydrosiloxane, polyvinylpyrrolidone-co-vinyl acetate, poly-dl-lactic acid, and poly l-lactic acid. It was found that none of the models were able to adequately predict in entirety the resultant dynamics. Of the models studied, the most successful were the HS upper bound (HSUB), the complementary universal Weiner equation (CWE), and the Lichtenecker model for the dimensional parameter, ζ = -1/2. The least successful models were the upper Weiner bound (UWB), the Neelakantaswamy, Turkman, and Sarkar (NTS) model for ζ = 1/2, and the Lichtenecker model for ζ = 1/2.
