Enhanced reporter gene expression in cells transfected in the presence of DMI-2, an acid nuclease inhibitor

Open Access
Publisher Website
Google Scholar
Abstract
Cellular nuclease activity is a potential barrier to the successful delivery of foreign genes to mammalian cells. We tested the hypothesis that transfection in the presence of a specific DNase inhibitor can enhance the expression of foreign gene products. We have used DMI-2, a polyketide metabolite of Streptomyces sp. strain 560 to enhance the expression of bacterial chloramphenicol acetyltransferase (CAT) in the human lung adenocarcinoma cell line H441. DMI-2 has been shown previously to inhibit porcine DNase II, an acid pH nuclease contained in the endosomal/lyso- somal compartment. Transfection of H441 cells in the presence of 0.1–1 μg/ml DMI-2 caused: (1) 10-fold enhancement of CAT activity when the bacterial plasmid was complexed with either surfactant protein A-poly-lysine or transferrin-poly-lysine; (2) 1.5- to two-fold enhancement of CAT activity in cells exposed to lipofectin–DNA complexes; (3) no effect on transfection via calcium phosphate co-precipitation. DMI-2 alone showed no inherent transfection activity. In experiments using SP-A-poly-lysine and plasmid containing the β-galactosidase reporter gene, DMI-2 increased the number of transfected cells. Methanolysis products of DMI-2 did not inhibit DNase II and did not enhance transfection efficiency. Taken together, the data support the hypothesis that nuclease action is a significant barrier to expression of foreign genes and inhibition of specific nucleases may facilitate transfection.

This publication has 0 references indexed in Scilit: