Ras-induced reactive oxygen species promote growth factor–independent proliferation in human CD34+ hematopoietic progenitor cells

Abstract

Excessive production of reactive oxygen species (ROS) is a feature of human malignancy and is often triggered by activation of oncogenes such as activated Ras. ROS act as second messengers and can influence a variety of cellular process including growth factor responses and cell survival. We have examined the contribution of ROS production to the effects of N-Ras G12D and H-Ras G12V on normal human CD34 ؉ progenitor cells. Activated Ras strongly up-regulated the production of both superoxide and hydrogen peroxide through the stimulation of NADPH oxidase (NOX) activity, without affecting the expression of endogenous antioxidants or the production of mitochondrially derived ROS. Activated Ras also promoted both the survival and the growth factor-independent proliferation of CD34 ؉ cells. Using oxidase inhibitors and antioxidants, we found that excessive ROS production by these cells did not contribute to their enhanced survival; rather, ROS promoted their growth factor-independent proliferation. Although Ras-induced ROS production specifically activated the p38 MAPK IntroductionReactive oxygen species (ROS) are a heterogeneous group of inorganic molecules and free radicals, with a wide spectrum of life span and reactivity. In physiologic systems, ROS formation begins with the univalent reduction of diatomic oxygen to produce superoxide radicals. There are 2 main sources of cellular superoxide; the first is the mitochondrial electron transport chain, where incomplete reduction of oxygen to water can result in superoxide formation. The other major source of superoxide is professional oxidases (exemplified by the NADPH oxidase [NOX] protein family), 1 which are expressed and functional throughout hematopoietic development. 2 These proteins form part of a membrane-bound complex that transfers single electrons from cytosolic nicotinamide adenine dinucleotide phosphate via flavin adenine dinucleotide to extracellular oxygen, producing superoxide. Superoxide is a shortlived radical but can dismutate forming hydrogen peroxide (H 2 O 2 ), a relatively long-lived species that lies at the hub of a variety of potential chemical reactions and is the main molecule from which all other physiologic ROS molecules are derived. ROS levels are regulated by a complex network of antioxidant molecules and enzymes that detoxify ROS. 3 For example, superoxide generation is balanced by the actions of superoxide dismutases (SODs), which convert superoxide to H 2 O 2 . Subsequently, H 2 O 2 is destroyed during oxidation reactions involving glutathione or members of the peroxiredoxin family.Although ROS production can have adverse consequences causing lipid peroxidation and DNA damage, it is also clear that ROS act as cell-signaling molecules. In recent years, it has become clear that H 2 O 2 reacts with a variety of proteins sensitive to thiol oxidation. 4 In particular, H 2 O 2 can inhibit phosphatases via oxidation of cysteine at the active site and may consolidate growth factor signaling by preventing futile cycles of phosphorylation a...

NOX-driven ROS formation in cell transformation of FLT3-ITD-positive AML

“…For example, early reports described enhanced ROS production in RAS-transformed fibroblasts [9], a finding which has later been corroborated in human CD34-positive hematopoietic progenitor cells [10]. More recently, also enhanced antioxidant activities were noted in cancer cells, for example as consequence of RAS-mediated transformation [11].…”

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“…[4][5][6] But what if cancer cells could facilitate this process too, by producing hydrogen peroxide themselves? In fact, oncogeneinduced transformation of cells results in hydrogen peroxide and ROS production [7][8][9][10] ( Fig. 1).…”

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