{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T19:40:55Z","timestamp":1760125255907,"version":"build-2065373602"},"reference-count":54,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2023,3,19]],"date-time":"2023-03-19T00:00:00Z","timestamp":1679184000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Institutes of Health","award":["GM128143","610311"],"award-info":[{"award-number":["GM128143","610311"]}]},{"name":"University of Minnesota","award":["GM128143","610311"],"award-info":[{"award-number":["GM128143","610311"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cells"],"abstract":"<jats:p>Nrf2 is the dominant cellular stress response factor that protects cells through transcriptional responses to xenobiotic and oxidative stimuli. Nrf2 malfunction is highly correlated with many human diseases, but the underlying molecular mechanisms remain to be fully uncovered. GATA4 is a conserved GATA family transcription factor that is essential for cardiac and dorsal epidermal development. Here, we describe a novel interaction between Drosophila Nrf2 and GATA4 proteins, i.e., cap\u2018n\u2019collar C (CncC) and Pannier (Pnr), respectively. Using the bimolecular fluorescence complementation (BiFC) assay\u2014a unique imaging tool for probing protein complexes in living cells\u2014we detected CncC\u2013Pnr complexes in the nuclei of Drosophila embryonic and salivary gland cells. Visualization of CncC\u2013Pnr BiFC signals on the polytene chromosome revealed that CncC and Pnr tend to form complexes in euchromatic regions, with a preference for loci that are not highly occupied by CncC or Pnr alone. Most genes within these loci are activated by the CncC\u2013Pnr BiFC, but not by individually expressed CncC or Pnr fusion proteins, indicating a novel mechanism whereby CncC and Pnr interact at specific genomic loci and coactivate genes at these loci. Finally, CncC-induced early lethality can be rescued by Pnr depletion, suggesting that CncC and Pnr function in the same genetic pathway during the early development of Drosophila. Taken together, these results elucidate a novel crosstalk between the Nrf2 xenobiotic\/oxidative response factor and GATA factors in the transcriptional regulation of development. This study also demonstrates that the polytene chromosome BiFC assay is a valuable tool for mapping genes that are targeted by specific transcription factor complexes.<\/jats:p>","DOI":"10.3390\/cells12060938","type":"journal-article","created":{"date-parts":[[2023,3,20]],"date-time":"2023-03-20T06:48:35Z","timestamp":1679294915000},"page":"938","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Determination of Complex Formation between Drosophila Nrf2 and GATA4 Factors at Selective Chromatin Loci Demonstrates Transcription Coactivation"],"prefix":"10.3390","volume":"12","author":[{"given":"Emma","family":"Neidviecky","sequence":"first","affiliation":[{"name":"Department of Biology, University of Minnesota Duluth, 1035 Kirby Drive, Duluth, MN 55812, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5867-3572","authenticated-orcid":false,"given":"Huai","family":"Deng","sequence":"additional","affiliation":[{"name":"Department of Biology, University of Minnesota Duluth, 1035 Kirby Drive, Duluth, MN 55812, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2023,3,19]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"76","DOI":"10.1101\/gad.13.1.76","article-title":"Keap1 represses nuclear activation of antioxidant responsive elements by Nrf2 through binding to the amino-terminal Neh2 domain","volume":"13","author":"Itoh","year":"1999","journal-title":"Genes Dev."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"273","DOI":"10.1007\/s00204-011-0675-4","article-title":"Nrf2: Control of sensitivity to carcinogens","volume":"85","author":"Slocum","year":"2011","journal-title":"Arch. Toxicol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"5718","DOI":"10.1093\/nar\/gkq212","article-title":"Global mapping of binding sites for Nrf2 identifies novel targets in cell survival response through ChIP-Seq profiling and network analysis","volume":"38","author":"Malhotra","year":"2010","journal-title":"Nucleic Acids Res."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"123","DOI":"10.1111\/j.1365-2443.2010.01473.x","article-title":"Molecular mechanisms of the Keap1-Nrf2 pathway in stress response and cancer evolution","volume":"16","author":"Taguchi","year":"2011","journal-title":"Genes Cells"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"769","DOI":"10.1080\/03602530600971974","article-title":"Mechanistic Studies of the Nrf2-Keap1 Signaling Pathway","volume":"38","author":"Zhang","year":"2006","journal-title":"Drug Metab. Rev."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"10070","DOI":"10.1073\/pnas.0502402102","article-title":"Modifying specific cysteines of the electrophile-sensing human Keap1 protein is insufficient to disrupt binding to the Nrf2 domain Neh2","volume":"102","author":"Eggler","year":"2005","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"85","DOI":"10.3389\/fonc.2017.00085","article-title":"The KEAP1\u2013NRF2 System in Cancer","volume":"7","author":"Taguchi","year":"2017","journal-title":"Front. Oncol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"51","DOI":"10.1007\/978-3-030-44599-7_3","article-title":"Nrf2 and the Nrf2-Interacting Network in Respiratory Inflammation and Diseases","volume":"85","author":"Carlson","year":"2020","journal-title":"Nrf2 Its Modul. Inflamm."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"e00467-19","DOI":"10.1128\/MCB.00467-19","article-title":"Nrf2 Suppresses Oxidative Stress and Inflammation in App Knock-In Alzheimer\u2019s Disease Model Mice","volume":"40","author":"Uruno","year":"2020","journal-title":"Mol. Cell. Biol."},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Smith, R.E., Tran, K., Smith, C.C., McDonald, M., Shejwalkar, P., and Hara, K. (2016). The Role of the Nrf2\/ARE Antioxidant System in Preventing Cardiovascular Diseases. Diseases, 4.","DOI":"10.3390\/diseases4040034"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"7","DOI":"10.4161\/fly.27007","article-title":"Multiple roles of Nrf2-Keap1 signaling","volume":"8","author":"Deng","year":"2013","journal-title":"Fly"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"302","DOI":"10.1016\/j.freeradbiomed.2015.06.020","article-title":"Mechanisms and functions of Nrf2 signaling in Drosophila","volume":"88","author":"Pitoniak","year":"2015","journal-title":"Free. Radic. Biol. Med."},{"key":"ref_13","first-page":"G1211","article-title":"Transcription factor Nrf2 regulates SHP and lipogenic gene expression in hepatic lipid metabolism","volume":"299","author":"Huang","year":"2010","journal-title":"Am. J. Physiol. Liver Physiol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"9292","DOI":"10.1074\/jbc.M109.093955","article-title":"Deficiency in the Nuclear Factor E2-related Factor-2 Transcription Factor Results in Impaired Adipogenesis and Protects against Diet-induced Obesity","volume":"285","author":"Pi","year":"2010","journal-title":"J. Biol. Chem."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"232","DOI":"10.1016\/j.stem.2016.04.015","article-title":"Mitochondrial Dynamics Impacts Stem Cell Identity and Fate Decisions by Regulating a Nuclear Transcriptional Program","volume":"19","author":"Khacho","year":"2016","journal-title":"Cell Stem Cell"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"66","DOI":"10.1016\/j.ccr.2012.05.016","article-title":"Nrf2 Redirects Glucose and Glutamine into Anabolic Pathways in Metabolic Reprogramming","volume":"22","author":"Mitsuishi","year":"2012","journal-title":"Cancer Cell"},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Deng, H., and Kerppola, T.K. (2013). Regulation of Drosophila Metamorphosis by Xenobiotic Response Regulators. PLoS Genet., 9.","DOI":"10.1371\/journal.pgen.1003263"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"109466","DOI":"10.1016\/j.celrep.2021.109466","article-title":"The Nrf2-Keap1 pathway is activated by steroid hormone signaling to govern neuronal remodeling","volume":"36","author":"Chew","year":"2021","journal-title":"Cell Rep."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"dev164384","DOI":"10.1242\/dev.164384","article-title":"GATA transcription factors in development and disease","volume":"145","author":"Tremblay","year":"2018","journal-title":"Development"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"107","DOI":"10.1016\/j.semcdb.2004.10.005","article-title":"GATA factors in Drosophila heart and blood cell development","volume":"16","author":"Sorrentino","year":"2005","journal-title":"Semin. Cell Dev. Biol."},{"key":"ref_21","first-page":"151","article-title":"Combined Mutation Screening of NKX2-5, GATA4, and TBX5 in Congenital Heart Disease: Multiple Heterozygosity and Novel Mutations","volume":"7","author":"Pope","year":"2011","journal-title":"Congenit. Hear. Dis."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"443","DOI":"10.1038\/nature01827","article-title":"GATA4 mutations cause human congenital heart defects and reveal an interaction with TBX","volume":"424","author":"Garg","year":"2003","journal-title":"Nature"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"5687","DOI":"10.1093\/emboj\/16.18.5687","article-title":"The cardiac transcription factors Nkx2-5 and GATA-4 are mutual cofactors","volume":"16","author":"Durocher","year":"1997","journal-title":"EMBO J."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"4611","DOI":"10.1038\/s41598-018-22830-8","article-title":"Cardiac Actions of a Small Molecule Inhibitor Targeting GATA4\u2013NKX2-5 Interaction","volume":"8","author":"Kinnunen","year":"2018","journal-title":"Sci. Rep."},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Kinnunen, S., V\u00e4lim\u00e4ki, M., T\u00f6lli, M., Wohlfahrt, G., Darwich, R., Komati, H., Nemer, M., and Ruskoaho, H. (2015). Nuclear Receptor-Like Structure and Interaction of Congenital Heart Disease-Associated Factors GATA4 and NKX2-5. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0144145"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"5679","DOI":"10.1242\/dev.126.24.5679","article-title":"The zinc finger proteins Pannier and GATA4 function as cardiogenic factors in Drosophila","volume":"126","author":"Gajewski","year":"1999","journal-title":"Development"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1277","DOI":"10.1242\/dev.119.4.1277","article-title":"Pannier, a negative regulator of achaete and scute in Drosophila, encodes a zinc finger protein with homology to the vertebrate transcription factor GATA-1","volume":"119","author":"Ramain","year":"1993","journal-title":"Development"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"7348","DOI":"10.1073\/pnas.97.13.7348","article-title":"The multitype zinc-finger protein U-shaped functions in heart cell specification in the Drosophila embryo","volume":"97","author":"Fossett","year":"2000","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"13174","DOI":"10.1038\/s41598-017-13487-w","article-title":"Angiotensin-converting enzyme Ance is cooperatively regulated by Mad and Pannier in Drosophila imaginal discs","volume":"7","author":"Kim","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1924","DOI":"10.1093\/molbev\/msaa335","article-title":"Variation in Pleiotropic Hub Gene Expression Is Associated with Interspecific Differences in Head Shape and Eye Size in Drosophila","volume":"38","author":"Buchberger","year":"2021","journal-title":"Mol. Biol. Evol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"258","DOI":"10.1016\/j.ydbio.2010.07.030","article-title":"Dorsal eye selector pannier (pnr) suppresses the eye fate to define dorsal margin of the Drosophila eye","volume":"346","author":"Oros","year":"2010","journal-title":"Dev. Biol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"3027","DOI":"10.1242\/dev.00517","article-title":"Gata factor Pannier is required to establish competence for heart progenitor formation","volume":"130","author":"Klinedinst","year":"2003","journal-title":"Development"},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Lovato, T.L., Sensibaugh, C.A., Swingle, K.L., Martinez, M.M., and Cripps, R.M. (2015). The Drosophila Transcription Factors Tinman and Pannier Activate and Collaborate with Myocyte Enhancer Factor-2 to Promote Heart Cell Fate. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0132965"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"86","DOI":"10.1002\/dvdy.22495","article-title":"Tinman is a direct activator of midline in the drosophila dorsal vessel","volume":"240","author":"Ryu","year":"2010","journal-title":"Dev. Dyn."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"3525","DOI":"10.1242\/dev.01899","article-title":"Hand is a direct target of Tinman and GATA factors during Drosophila cardiogenesis and hematopoiesis","volume":"132","author":"Han","year":"2005","journal-title":"Development"},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Jin, H., Stojnic, R., Adryan, B., Ozdemir, A., Stathopoulos, A., and Frasch, M. (2013). Genome-Wide Screens for In Vivo Tinman Binding Sites Identify Cardiac Enhancers with Diverse Functional Architectures. PLoS Genet., 9.","DOI":"10.1371\/journal.pgen.1003195"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"719","DOI":"10.1242\/dev.118.3.719","article-title":"The gene tinman is required for specification of the heart and visceral muscles in Drosophila","volume":"118","author":"Bodmer","year":"1993","journal-title":"Development"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"251","DOI":"10.1016\/j.bbaexp.2004.01.002","article-title":"Regulation of human microsomal epoxide hydrolase gene (EPHX1) expression by the transcription factor GATA-4","volume":"1676","author":"Zhu","year":"2004","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"863","DOI":"10.1016\/j.ibmb.2011.07.002","article-title":"Induction of a detoxification gene in Drosophila melanogaster requires an interaction between tissue specific enhancers and a novel cis-regulatory element","volume":"41","author":"Chung","year":"2011","journal-title":"Insect Biochem. Mol. Biol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"103653","DOI":"10.1016\/j.etap.2021.103653","article-title":"Ambient air PM2.5 exposure induces heart injury and cardiac hypertrophy in rats through regulation of miR-208a\/b, \u03b1\/\u03b2-MHC, and GATAEnviron","volume":"85","author":"Bai","year":"2021","journal-title":"Toxicol. Pharmacol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"789","DOI":"10.1016\/S1097-2765(02)00496-3","article-title":"Visualization of Interactions among bZIP and Rel Family Proteins in Living Cells Using Bimolecular Fluorescence Complementation","volume":"9","author":"Hu","year":"2002","journal-title":"Mol. Cell"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"76","DOI":"10.1016\/j.devcel.2007.12.002","article-title":"Keap1\/Nrf2 Signaling Regulates Oxidative Stress Tolerance and Lifespan in Drosophila","volume":"14","author":"Sykiotis","year":"2008","journal-title":"Dev. Cell"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"4023","DOI":"10.1242\/dev.127.18.4023","article-title":"Cap \u2018n\u2019 collar B cooperates with a small Maf subunit to specify pharyngeal development and suppress deformed homeotic function in the Drosophila head","volume":"127","author":"Veraksa","year":"2000","journal-title":"Development"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"3277","DOI":"10.1242\/dev.110528","article-title":"Visualization of the Drosophila dKeap1-CncC interaction on chromatin illumines cooperative, xenobiotic-specific gene activation","volume":"141","author":"Deng","year":"2014","journal-title":"Development"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"429","DOI":"10.1016\/bs.mie.2017.02.003","article-title":"Visualization of the Genomic Loci That Are Bound by Specific Multiprotein Complexes by Bimolecular Fluorescence Complementation Analysis on Drosophila Polytene Chromosomes","volume":"589","author":"Deng","year":"2017","journal-title":"Methods Enzymol."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"387","DOI":"10.1016\/j.ymeth.2009.02.019","article-title":"Polytene chromosome squash methods for studying transcription and epigenetic chromatin modification in Drosophila using antibodies","volume":"48","author":"Johansen","year":"2009","journal-title":"Methods"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"175","DOI":"10.1016\/S1097-2765(03)00011-X","article-title":"A Cell Cycle-Regulated GATA Factor Promotes Centromeric Localization of CENP-A in Fission Yeast","volume":"11","author":"Chen","year":"2003","journal-title":"Mol. Cell"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"1796","DOI":"10.1101\/gad.17280911","article-title":"Transcriptional regulation of xenobiotic detoxification in Drosophila","volume":"25","author":"Misra","year":"2011","journal-title":"Genes Dev."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"e27","DOI":"10.1093\/nar\/gkx1246","article-title":"TAD-free analysis of architectural proteins and insulators","volume":"46","author":"Mourad","year":"2017","journal-title":"Nucleic Acids Res."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"473","DOI":"10.1016\/j.cell.2012.01.030","article-title":"A Transcription Factor Collective Defines Cardiac Cell Fate and Reflects Lineage History","volume":"148","author":"Junion","year":"2012","journal-title":"Cell"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"21","DOI":"10.1016\/j.ccell.2018.03.022","article-title":"NRF2 and the Hallmarks of Cancer","volume":"34","author":"Chapman","year":"2018","journal-title":"Cancer Cell"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"4553","DOI":"10.1242\/dev.125.22.4553","article-title":"A cap \u2018n\u2019 collar protein isoform contains a selective Hox repressor function","volume":"125","author":"McGinnis","year":"1998","journal-title":"Development"},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"356","DOI":"10.1016\/j.ydbio.2005.12.022","article-title":"Modulation of AP and DV signaling pathways by the homeotic gene Ultrabithorax during haltere development in Drosophila","volume":"291","author":"Mohit","year":"2006","journal-title":"Dev. Biol."},{"key":"ref_54","doi-asserted-by":"crossref","unstructured":"Deng, H. (2020). Nrf2 and Its Modulation in Inflammation, Springer International Publishing.","DOI":"10.1007\/978-3-030-44599-7"}],"container-title":["Cells"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4409\/12\/6\/938\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T18:58:50Z","timestamp":1760122730000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4409\/12\/6\/938"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,3,19]]},"references-count":54,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2023,3]]}},"alternative-id":["cells12060938"],"URL":"https:\/\/doi.org\/10.3390\/cells12060938","relation":{},"ISSN":["2073-4409"],"issn-type":[{"type":"electronic","value":"2073-4409"}],"subject":[],"published":{"date-parts":[[2023,3,19]]}}}