PLANT NITRATE REDUCTASE GENE FRAGMENTS ENHANCE NITRITE PRODUCTION IN ACTIVATED MURINE MACROPHAGE CELL-LINES

Nitrate reductase (NR) gene fragments (1.1 kb and 800 bp) from the bar ley plant were incorporated into pSV(2)neo and transfected by electrop oration into a variety of cell lines of different functionality. Only transfected murine macrophage cell lines demonstrated appreciably enha nced NO2- production (i.e., NR activity) both in the presence and abse nce of exogenous nitrate (NO3-). Addition of NO3- caused the greatest increase in NO2- production when macrophages were primed with interfer on-gamma (INF-gamma) and lipopolysaccharide (LPS). Transfection of RAW 264.7 murine macrophages led to isolation of several novel neomycin-r esistant subpopulations designated NR10(1), NR10(2) (both containing t he 1.1 kb NR fragment) and NR800(5) (containing the 800 bp NR fragment ). Similarly transfected nonleukocytic and leukocytic stem cell lines showed no significant NO2- production. Outside of the macrophage cell lines, only the murine T cell line EL-4 showed evidence of mild nitrit e production enhancement. The mechanism of enhanced NO2- formation in NR transfected murine macrophages is unknown. However, study of these novel cells may lead to greater understanding of the expression of a p lant NR in mammalian cells and highly controlled production of a cytot oxic molecule (NO2-) in macrophages. (C) 1994 Academic Press, Inc.