EglN2 associates with the NRF1 ‐ PGC1α complex and controls mitochondrial function in breast cancer by Jing Zhang, Chengyang Wang, Xi Chen, Mamoru Takada, Cheng Fan, Xingnan Zheng, Haitao Wen, Yong Liu, Chenguang Wang, Richard G Pestell, Katherine M Aird, William G Kaelin, Xiaole Shirley Liu, and Qing Zhang EMBO J. Volume ():e October 22, 2015 ©2015 by European Molecular Biology Organization

EglN2 affects mitochondrial function in ERα ‐ positive breast cancer cells A, BImmunoblot from T47D (A) or MCF ‐ 7 (B) cells infected with lentivirus encoding EglN2 shRNA (326, 327 or 328) or control shRNA (Ctrl).C, DMeasurement of oxygen consumption rate (OCR... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

EglN2 regulates mitochondrial function A–CImmunoblot of cell lysates (A), measurement of oxygen consumption rate (B), and qRT–PCR quantification of mtDNA (C) from T47D cells infected with lentivirus encoding either EglN2 shRNA or control (Ctrl) shRNA follo... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

EglN2 regulates mitochondrial function in a HIF1/2α ‐ independent manner A, BImmunoblot from T47D (A) or MCF ‐ 7 (B) cells infected with lentivirus encoding EglN2 shRNA (326, 327, or 328) or control shRNA (Ctrl).C–HImmunoblot of cell lysates (C, E, G) and qRT–... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

EglN2 regulates mitochondrial function in a HIF1/2α ‐ independent manner A–CMeasurement of oxygen consumption from MCF7 cells infected with lentivirus encoding either HIF1 shRNAs (3809, 3810) (A), HIF2 shRNA (3804) (B), ARNT shRNA (1770) (C), or control (Ctr... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

EglN2 binds to chromatin and regulates mitochondrial function under hypoxia A, BqRT–PCR quantification of mtDNA from T47D (A) or MCF ‐ 7 (B) cells infected with lentivirus encoding EglN2 shRNA (326, 327, or 328) or control shRNA and treated with hypoxia (5%... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

EglN2 binds to chromatin and regulates mitochondrial function under hypoxia AMeasurement of oxygen consumption by an oxytherm electrode unit as a function of time for MCF7 cells infected with lentivirus encoding EglN2 shRNA (327 or 328) or control shRNA tr... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

EglN2 interacts with NRF1 and PGC1α AGenomewide distribution of EglN2 binding peaks under hypoxia (1% O2) with concurrent DMOG (1 mM) treatment for 16 h.BMotif enrichment of integrated analyses of gene expression profiling (Ctrl vs EglN2) and EglN2 binding... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

EglN2 binds to NRF1 and PGC1α complex on chromatin A, BAnalysis and comparison of gene expression profiling (siCtrl vs siEglN2) between normoxia and hypoxia (1% O2) in T47D.CRepresentative images (left) and quantification (right) of anchorage ‐ independent g... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

EglN2 and NRF1 regulate FDXR in ERα ‐ positive breast cancer cells AGenomewide distribution of NRF1 binding peaks under hypoxia (1% O2) with concurrent DMOG (1 mM) treatment for 16 h.BVenn diagram showing the overlap between the EglN2 and NRF1 cistromes in T... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

EglN2/NRF1/PGC1α regulates FDXR in breast cancer A, BqRT–PCR of mRNA from T47D transfected with either EglN2 siRNA (1, 4) (A) or NRF1 pool siRNA (B) under normoxia.CqRT– PCR of mRNA from T47D with or without hypoxia (5% O2) treatment.DqRT–PCR of mRNA from T... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization

FDXR regulates mitochondrial function and ERα ‐ positive breast tumorigenesis A, BqRT–PCR of mRNA (A) and mitochondrial DNA (B) from T47D cells infected with lentivirus encoding either FDXR shRNA (434, 435) or control (Ctrl) shRNA followed by hypoxia treatme... Jing Zhang et al. EMBO J. doi: /embj ©2015 by European Molecular Biology Organization