Superoxide generation in v-Ha-ras-transduced human keratinocyte HaCaT cells

Abstract

The oncogenic ras protein controls signal-transduction pathways that are critical for cell proliferation and tumorigenesis. Here, we demonstrate that v-Ha-ras-transduced human keratinocyte HaCaT cells produced significantly larger amounts of superoxide than did control cell lines. The superoxide generation was mediated by the transduced ras protein, because superoxide generation was modified by an inhibitor, lovastatin, that inhibits ras farnesylation during ras protein maturation. Superoxide generation was also inhibited by diphenylene iodonium, an inhibitor of flavoproteins, including NADPH oxidase, but not by rotenone, an inhibitor of the respiratory chain of the mitochondria. Those observations suggested that a phagocytic-like NADPH oxidase exists in keratinocytes that could be activated by the dominant activated v-Ha-ras and produce superoxide. Overexpression of manganese-containing superoxide dismutase and copper and zinc-containing superoxide dismutase cDNA via adenovirus infection also attenuated superoxide generation. Previous work has demonstrated that extracellular superoxide dismutase (SOD) can lower superoxide generation; this is the first report that intracellular SOD could also modify the amount of superoxide production from the cells. This report implies that superoxide radical may act as a second messenger molecule of oncogenic ras.

Role of Phagocyte Oxidase in UVA-Induced Oxidative Stress and Apoptosis in Keratinocytes

“…gp91phox (NOX2), the founding member of the phagocyte NADPH oxidase family, is predominantly expressed in phagocytes such as neutrophils and plays a crucial role in host defense against microbial infection. Recently NADPH oxidase has been shown to be involved in the generation of superoxide (Yang et al, 1999), DNA repair (Cho et al, 2002), wound healing (Sen et al, 2002), and activation of p38 (Chiu et al, 2001) in keratinocytes. The role of one member gp91phox in UVA-exposed keratinocytes, however, is not known.…”

l-NAME has Opposite Effects on the Productions of S-adenosylhomocysteine and S-adenosylmethionine in V12-H-Ras and M-CR3B-Ras Pheochromocytoma Cells

“…NO suppressed the prooxidant and proapoptotic effects of ceramide by maintaining intracellular iron homeostasis and inversed relationship between endogenous NO and cellular iron levels. Ceramide-induced oxidative stress was thought to result activation of Rac 1 [28], which through NADPH oxidase/ROS control a variety of Ras-mediated cellular effects, including cell transformation [29]. Thus, endogenous NO through iron flow and activity of cystathionine b-synthase may regulate methyl cycle.…”

Differential induction of reactive oxygen species through Erk1/2 and Nox-1 by FK228 for selective apoptosis of oncogenic H-Ras-expressing human urinary bladder cancer J82 cells

“…To determine the role of Nox-1 in FK228-induced diVerential increases of ROS and cell death in J82-Ras cells versus J82 cells, we initially used DPI, which is a Xavoprotein inhibitor (Yang et al 1999;Riganti et al 2004), to suppress NADPH oxidases. We detected that DPI treatment signiWcantly reduced FK228-induced ROS production ( Fig.…”

“…Elevation of ROS has been shown to play an important role in Ras-induced transformation phenotypes (Cho et al 2002;Yang et al 1999;Santillo et al 2001;Liu et al 2001;Mitsushita et al 2004;Shinohara et al 2007). Expression of oncogenic H-Ras results in elevating intracellular ROS through the ERK pathway to up-regulate NADPH oxidase-1 (Nox-1) gene expression (Adachi et al 2008).…”