Mitochondrial reactive oxygen species mediate GPCR-induced TACE/ADAM17-dependent transforming growth factor-alpha shedding.
Epidermal growth factor receptor (EGFR) activation by GPCRs regulates many important biological processes. ADAM metalloprotease activity has been implicated as a key step in transactivation, yet the regulatory mechanisms are not fully understood. Here, we investigate the regulation of transforming growth factor-alpha (TGF-alpha) shedding by reactive oxygen species (ROS) through the ATP-dependent activation of the P2Y family of GPCRs. We report that ATP stimulates TGF-alpha proteolysis with concomitant EGFR activation and that this process requires TACE/ADAM17 activity in both murine fibroblasts and CHO cells. ATP-induced TGF-alpha shedding required calcium and was independent of Src family kinases and PKC and MAPK signaling. Moreover, ATP-induced TGF-alpha shedding was completely inhibited by scavengers of ROS, whereas calcium-stimulated shedding was partially inhibited by ROS scavenging. Hydrogen peroxide restored TGF-alpha shedding after calcium chelation. Importantly, we also found that ATP-induced shedding was independent of the cytoplasmic NADPH oxidase complex. Instead, mitochondrial ROS production increased in response to ATP and mitochondrial oxidative complex activity was required to activate TACE-dependent shedding. These results reveal an essential role for mitochondrial ROS in regulating GPCR-induced growth factor shedding.
Myers
Timothy J
TJ
Department of Biochemistry and Biophysics and Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA.
Brennaman
Leann H
LH
Stevenson
Mary
M
Higashiyama
Shigeki
S
Russell
William E
WE
Lee
David C
DC
Sunnarborg
Susan Wohler
SW
D051722:ADAM Proteins / Q000378:metabolism*; D000072198:ADAM17 Protein; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism; D018922:HL-60 Cells; D006801:Humans; D051379:Mice; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D019255:NADPH Oxidases / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D018048:Receptors, Purinergic P2 / Q000378:metabolism; D015533:Transcriptional Activation / Q000187:drug effects; D016211:Transforming Growth Factor alpha / Q000378:metabolism*
D017382:Reactive Oxygen Species; D043562:Receptors, G-Protein-Coupled; D018048:Receptors, Purinergic P2; D016211:Transforming Growth Factor alpha; D000255:Adenosine Triphosphate; D019255:NADPH Oxidases; D066246:ErbB Receptors; D051722:ADAM Proteins; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human; C000606463:Adam17 protein, mouse; D002118:Calcium