title: Subcellular localization of the FLT3-ITD oncogene plays a significant role in the production of NOX- and p22phox-derived reactive oxygen species in acute myeloid leukemia.
abstract: Internal tandem duplication of the juxtamembrane domain of FMS-like tyrosine kinase 3 (FLT3-ITD) receptor is the most prevalent FLT3 mutation accounting for 20% of acute myeloid leukemia (AML) patients. FLT3-ITD mutation results in ligand-independent constitutive activation of the receptor at the plasma membrane and 'impaired trafficking' of the receptor in compartments of the endomembrane system, such as the endoplasmic reticulum (ER). FLT3-ITD expressing cells have been shown to generate increased levels of reactive oxygen species (ROS), in particular NADPH oxidase (NOX)-generated ROS which act as pro-survival signals. The purpose of this study is to investigate FLT3-ITD production of ROS at the plasma membrane and ER in the FLT3-ITD expressing AML cell line MV4-11. Receptor trafficking inhibitors; Tunicamycin and Brefeldin A induce ER retention of FLT3-ITD, resulting in a decrease in protein expression of NOX4 and its partner protein p22phox, thus demonstrating the critical importance of FLT3-ITD localization for the generation of pro-survival ROS. NOX-generated ROS contribute to total endogenous hydrogen peroxide (H2O2) in AML as quantified by flow cytometry using the cell-permeable H2O2-probe Peroxy Orange 1 (PO1). We found that PI3K/AKT signaling only occurs when FLT3-ITD is expressed at the plasma membrane and is required for the production of NOX-generated ROS. ER retention of FLT3-ITD resulted in NOX4 deglycosylation and p22phoxprotein degradation.
authors.affiliation: Tumour Biology Laboratory, School of Biochemistry and Cell Biology, Bioscience Research Institute, University College Cork, Cork, Ireland.
authors.forename: Jennifer N
authors.initials: JN
authors.lastname: Moloney
authors.affiliation: Tumour Biology Laboratory, School of Biochemistry and Cell Biology, Bioscience Research Institute, University College Cork, Cork, Ireland.
authors.forename: Joanna
authors.initials: J
authors.lastname: Stanicka
authors.affiliation: Tumour Biology Laboratory, School of Biochemistry and Cell Biology, Bioscience Research Institute, University College Cork, Cork, Ireland. Electronic address: t.cotter@ucc.ie.
authors.forename: Thomas G
authors.initials: TG
authors.lastname: Cotter
mesh_terms: D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000235:genetics* / Q000378:metabolism; D019255:NADPH Oxidases / Q000378:metabolism*; D009857:Oncogenes; D017382:Reactive Oxygen Species / Q000378:metabolism*; D020080:Tandem Repeat Sequences; D051941:fms-Like Tyrosine Kinase 3 / Q000235:genetics* / Q000378:metabolism
chemical_list: D017382:Reactive Oxygen Species; D019255:NADPH Oxidases; C086986:CYBA protein, human; C497970:FLT3 protein, human; D051941:fms-Like Tyrosine Kinase 3
keywords: Acute myeloid leukemia; FLT3-ITD; NADPH oxidase; Oncogene; Pro-survival reactive oxygen species; p22(phox)