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.

Superoxide Dismutases in Pancreatic Cancer

“…Growth factors were shown to stimulate O 2 ●− generation in pancreatic cancer cells through the activation of NOX, whereas inhibition of O 2 ●− production induced apoptosis. Similar results have been found in K- ras transformed human keratinocytes [ 42 ]. The increased levels of O 2 ●− could be reversed efficiently by intracellular SODs.…”

Regulation of pancreatic cancer growth by superoxide

“…Based on these observations it is hypothesized that K- ras may activate the NADPH oxidase (NOX) system to produce O 2 ·− that leads to cell proliferation. Similar results have been found in human keratinocytes (5). In ras transformed keratinocytes, increased O 2 ·− production was demonstrated and this increased production could be blocked efficiently by superoxide dismutase (SOD).…”

The Interplay between ROS and Ras GTPases: Physiological and Pathological Implications

“…In the same study experimental evidence was provided suggesting that Rac is positioned downstream to Ras. Similar overexpression of Ras in other cell types such as keratinocytes [40] and epithelial cells [41] also demonstrated an increase in basal ROS levels. The pathway by which Ras regulates the levels of ROS remains incompletely understood.…”