Co-transfection of 293 cells with siRNA and plasmid DNA using DreamFect Gold

293 cells were co-transfected with siRNA and plasmid vectors using DreamFect Gold transfection reagent.

This article demonstrates that due to its low toxicity level and to its high transfection capacities, DreamFect Gold from OZ Biosciences is the reagent of choice for co-transfecting cell lines.

article reference: Biochim Biophys Acta. 2013 Jul 21. pii: S0304-4165(13)00320-6.

Hypoxia inducible NOD2 interacts with 3-O-sulfogalactoceramide and regulates vesicular homeostasis.
Nabatov AA, Hatzis P, Rouschop KM, van Diest P, Vooijs M.

Abstract

BACKGROUND: Oxygen sensing in mammalian cells is a conserved signaling pathway regulated by hypoxia inducible factor type 1 (HIF-1). Inadequate oxygen supply (hypoxia) is common to many pathological disorders where autophagy plays an import role. The aim of this study was the identification and characterization of novel HIF-1 target genes that promote autophagy during hypoxia.
METHODS: Whole genome Chromatin Immune Precipitation from hypoxic HeLa cells was used to identify novel HIF-1 target genes. Hypoxia induced expression and transcription regulation was studied in wild type and HIF-deficient cells. siRNA silencing of candidate genes was used to establish their role during autophagy. Recombinant protein was used for screening immobilized glycosylated lipids to identify potential ligands.
RESULTS: We identified the Nucleotide Oligomerization Domain 2 (NOD2/CARD15) as a novel HIF-1 target and 3-O-sulfo-galactoceramide (sulfatide) and Mycobacterium sp. specific sulfolipid-1 as the first NOD2 ligands that both compete for binding to NOD2. Loss of NOD2 function impaired autophagy upstream of the autophagy inhibitor chloroquine by reducing the number of acidic vesicles. Inhibition of sulfatide synthesis elicited defects in autophagy similar to the NOD2 loss of function but did not influence NOD2-mediated NF-kB signaling.
CONCLUSIONS: Our findings suggest that the interaction of NOD2 with sulfatide may mediate the balance between autophagy and inflammation in hypoxic cells.
GENERAL SIGNIFICANCE: These findings may lead to a better understanding of complex inflammatory pathologies like Crohn's disease and tuberculosis where both NOD2 and hypoxia are implicated.

DreamFect™Gold allows transfecting all types of nucleic acids with a very high efficiency. Due to its formulation, DreamFect™Gold delivers a large quantity of nucleic acids leading to higher protein expression compared to other transfection reagents. It is fully biodegradable and does not interfere with cellular mechanisms.