Liposome-based DNA carriers may induce cellular stress response and change gene expression pattern in transfected cells

Abstract

BackgroundDuring functional studies on the rat stress-inducible Hspa1b (hsp70.1) gene we noticed that some liposome-based DNA carriers, which are used for transfection, induce its promoter activity. This observation concerned commercial liposome formulations (LA), Lipofectin and Lipofectamine 2000. This work was aimed to understand better the mechanism of this phenomenon and its potential biological and practical consequences.ResultsWe found that a reporter gene driven by Hspa1b promoter is activated both in the case of transient transfections and in the stably transfected cells treated with LA. Using several deletion clones containing different fragments of Hspa1b promoter, we found that the regulatory elements responsible for most efficient LA-driven inducibility were located between nucleotides -269 and +85, relative to the transcription start site. Further studies showed that the induction mechanism was independent of the classical HSE-HSF interaction that is responsible for gene activation during heat stress. Using DNA microarrays we also detected significant activation of the endogenous Hspa1b gene in cells treated with Lipofectamine 2000. Several other stress genes were also induced, along with numerous genes involved in cellular metabolism, cell cycle control and pro-apoptotic pathways.ConclusionsOur observations suggest that i) some cationic liposomes may not be suitable for functional studies on hsp promoters, ii) lipofection may cause unintended changes in global gene expression in the transfected cells.

DNA Damage Follows Repair Factor Depletion and Portends Genome Variation in Cancer Cells after Pore Migration

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Transfecting RAW264.7 Cells with a Luciferase Reporter Gene

“…Lipid-based transfection reagents are typically used in research due to its ease of use and commercial availability. However, some of these reagents caused unintended (and usually unwanted) changes in global gene expression in transfected cells 11,12 . To address this problem, cells are only incubated with the transfection reagents for a few hours instead of the typical O/N, or a non-lipid-based reagent is chosen for transfection.…”

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“…Transfection of cells can elicit a heat-shock response (e.g. 89), raising the possibility that chaperone proteins become limiting under these conditions. Thus, a role for chaperones in protecting activated PKCs from dephosphorylation together with the require-FIGURE 7.…”