{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T01:17:18Z","timestamp":1760231838478,"version":"build-2065373602"},"reference-count":46,"publisher":"MDPI AG","issue":"5","license":[{"start":{"date-parts":[[2022,5,21]],"date-time":"2022-05-21T00:00:00Z","timestamp":1653091200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000002","name":"NIH","doi-asserted-by":"publisher","award":["R01 GM105964","R21 NS125242"],"award-info":[{"award-number":["R01 GM105964","R21 NS125242"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biomolecules"],"abstract":"<jats:p>The human (h) transporter hZIP4 is the primary Zn2+ importer in the intestine. hZIP4 is also expressed in a variety of organs such as the pancreas and brain. Dysfunction of hZIP4 can result in the Zn2+ deficiency disease acrodermatitis enteropathica (AE). AE can disrupt digestive and immune system homeostasis. A limited number of hZIP4 expression strategies have hindered increasing knowledge about this essential transmembrane protein. Here, we report the heterologous expression of hZIP4 in Saccharomyces cerevisiae. Both a wild-type and a mutant S. cerevisiae strain, in which the endogenous Zn2+ transporters were deleted, were used to test the expression and localization of an hZIP4\u2013GFP fusion protein. A full-length hZIP4\u2013GFP and a truncated membrane-domain-only (mhZIP4\u2013GFP) protein were observed to be present in the plasma membrane in yeast.<\/jats:p>","DOI":"10.3390\/biom12050726","type":"journal-article","created":{"date-parts":[[2022,5,21]],"date-time":"2022-05-21T09:18:08Z","timestamp":1653124688000},"page":"726","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Heterologous Expression of Full-Length and Truncated Human ZIP4 Zinc Transporter in Saccharomyces cerevisiae"],"prefix":"10.3390","volume":"12","author":[{"given":"Yuting","family":"Liu","sequence":"first","affiliation":[{"name":"Department of Chemistry and Biochemistry, Worcester Polytechnic Institute, Worcester, MA 01609, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Elizabeth M.","family":"Bafaro","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, Worcester Polytechnic Institute, Worcester, MA 01609, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Robert E.","family":"Dempski","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry, Worcester Polytechnic Institute, Worcester, MA 01609, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2022,5,21]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"612","DOI":"10.1016\/j.mam.2012.05.011","article-title":"The SLC39 family of zinc transporters","volume":"34","author":"Jeong","year":"2013","journal-title":"Mol. Asp. Med."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"331","DOI":"10.1023\/A:1012905406548","article-title":"Functions of zinc in signaling, proliferation and differentiation of mammalian cells","volume":"14","author":"Beyersmann","year":"2001","journal-title":"BioMetals"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"17029","DOI":"10.1038\/sigtrans.2017.29","article-title":"The emerging role of zinc transporters in cellular homeostasis and cancer","volume":"2","author":"Bafaro","year":"2017","journal-title":"Signal Transduct. Target. Ther."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"176","DOI":"10.3945\/an.112.003210","article-title":"Discovery of Human Zinc Deficiency: Its Impact on Human Health and Disease","volume":"4","author":"Prasad","year":"2013","journal-title":"Adv. Nutr. Int. Rev. J."},{"key":"ref_5","first-page":"144","article-title":"Zinc and its importance for human health: An integrative review","volume":"18","author":"Roohani","year":"2013","journal-title":"J. Res. Med. Sci. Off. J. Isfahan Univ. Med. Sci."},{"key":"ref_6","unstructured":"Arg\u00fcello, J.M., and Lutsenko, S. (2012). Chapter Nine\u2013The Cation Selectivity of the ZIP Transporters. Current Topics in Membranes, Academic Press."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"18636","DOI":"10.1073\/pnas.0709307104","article-title":"Aberrant expression of zinc transporter ZIP4 (SLC39A4) significantly contributes to human pancreatic cancer pathogenesis and progression","volume":"104","author":"Li","year":"2007","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"33474","DOI":"10.1074\/jbc.M305000200","article-title":"The Acrodermatitis Enteropathica Gene ZIP4 Encodes a Tissue-specific, Zinc-regulated Zinc Transporter in Mice","volume":"278","author":"Wang","year":"2003","journal-title":"J. Biol. Chem."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1423","DOI":"10.1158\/1078-0432.CCR-09-2405","article-title":"ZIP4 Regulates Pancreatic Cancer Cell Growth by Activating IL-6\/STAT3 Pathway via Zinc Finger Transcription Factor CREB","volume":"16","author":"Zhang","year":"2010","journal-title":"Clin. Cancer Res. Off. J. Am. Assoc. Cancer Res."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"66","DOI":"10.1086\/341125","article-title":"A Novel Member of a Zinc Transporter Family Is Defective in Acrodermatitis Enteropathica","volume":"71","author":"Wang","year":"2002","journal-title":"Am. J. Hum. Genet."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"239","DOI":"10.1038\/ng913","article-title":"Identification of SLC39A4, a gene involved in acrodermatitis enteropathica","volume":"31","author":"Giraudet","year":"2002","journal-title":"Nat. Genet."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1301","DOI":"10.1515\/BC.2007.149","article-title":"Novel zinc-responsive post-transcriptional mechanisms reciprocally regulate expression of the mouse Slc39a4 and Slc39a5 zinc transporters (Zip4 and Zip5)","volume":"388","author":"Weaver","year":"2007","journal-title":"Biol. Chem."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1242","DOI":"10.1042\/BST0361242","article-title":"Regulation and Function of Zip4, the Acrodermatitis Enteropathica Gene","volume":"36","author":"Andrews","year":"2008","journal-title":"Biochem. Soc. Trans."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"129","DOI":"10.1128\/MCB.00963-08","article-title":"Novel Proteolytic Processing of the Ectodomain of the Zinc Transporter ZIP4 (SLC39A4) during Zinc Deficiency Is Inhibited by Acrodermatitis Enteropathica Mutations","volume":"29","author":"Kambe","year":"2009","journal-title":"Mol. Cell. Biol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"5376","DOI":"10.1158\/1078-0432.CCR-08-0455","article-title":"Overexpression of the zinc uptake transporter hZIP1 inhibits nuclear factor-kappaB and reduces the malignant potential of prostate cancer cells in vitro and in vivo","volume":"14","author":"Golovine","year":"2008","journal-title":"Clin. Cancer Res. Off. J. Am. Assoc. Cancer Res."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"5560","DOI":"10.1074\/jbc.275.8.5560","article-title":"Functional Expression of the Human hZIP2 Zinc Transporter","volume":"275","author":"Gaither","year":"2000","journal-title":"J. Biol. Chem."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"17796","DOI":"10.1074\/jbc.M114.617613","article-title":"Computation and Functional Studies Provide a Model for the Structure of the Zinc Transporter hZIP4","volume":"290","author":"Antala","year":"2015","journal-title":"J. Biol. Chem."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1705","DOI":"10.1021\/acs.biochem.9b00131","article-title":"Quantifying the Oligomeric State of hZIP4 on the Surface of Cells","volume":"58","author":"Ahern","year":"2019","journal-title":"Biochemistry"},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Hoch, E., Levy, M., Hershfinkel, M., and Sekler, I. (2020). Elucidating the H+ Coupled Zn2+ Transport Mechanism of ZIP4; Implications in Acrodermatitis Enteropathica. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21030734"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"868","DOI":"10.1002\/pro.3591","article-title":"Concomitant disorder and high-affinity zinc binding in the human zinc- and iron-regulated transport protein 4 intracellular loop","volume":"28","author":"Bafaro","year":"2019","journal-title":"Protein Sci."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"337","DOI":"10.1002\/humu.9178","article-title":"Mutation spectrum of human SLC39A4 in a panel of patients with acrodermatitis enteropathica","volume":"22","author":"Kharfi","year":"2003","journal-title":"Hum. Mutat."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"6992","DOI":"10.1074\/jbc.M610552200","article-title":"A Histidine-rich Cluster Mediates the Ubiquitination and Degradation of the Human Zinc Transporter, hZIP4, and Protects against Zinc Cytotoxicity","volume":"282","author":"Mao","year":"2007","journal-title":"J. Biol. Chem."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"963","DOI":"10.1021\/bi201553p","article-title":"The Human ZIP4 Transporter Has Two Distinct Binding Affinities and Mediates Transport of Multiple Transition Metals","volume":"51","author":"Antala","year":"2012","journal-title":"Biochemistry"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"4523","DOI":"10.1074\/jbc.M310799200","article-title":"Zn2+-stimulated Endocytosis of the mZIP4 Zinc Transporter Regulates Its Location at the Plasma Membrane","volume":"279","author":"Kim","year":"2004","journal-title":"J. Biol. Chem."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1319","DOI":"10.1039\/C5MT00066A","article-title":"The large intracellular loop of hZIP4 is an intrinsically disordered zinc binding domain","volume":"7","author":"Bafaro","year":"2015","journal-title":"Met. Integr. Biomet. Sci."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e1700344","DOI":"10.1126\/sciadv.1700344","article-title":"Crystal structures of a ZIP zinc transporter reveal a binuclear metal center in the transport pathway","volume":"3","author":"Zhang","year":"2017","journal-title":"Sci. Adv."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"785","DOI":"10.1007\/s10534-011-9451-4","article-title":"Transition metal homeostasis: From yeast to human disease","volume":"24","author":"Bleackley","year":"2011","journal-title":"BioMetals"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"26092","DOI":"10.1016\/S0021-9258(18)47163-3","article-title":"The FET4 gene encodes the low affinity Fe(II) transport protein of Saccharomyces cerevisiae","volume":"269","author":"Dix","year":"1994","journal-title":"J. Biol. Chem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"4112","DOI":"10.1074\/jbc.M807909200","article-title":"Copper Transport Activity of Yeast Ctr1 Is Down-regulated via Its C Terminus in Response to Excess Copper","volume":"284","author":"Wu","year":"2009","journal-title":"J. Biol. Chem."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"23203","DOI":"10.1074\/jbc.271.38.23203","article-title":"The ZRT2 Gene Encodes the Low Affinity Zinc Transporter in Saccharomyces cerevisiae","volume":"271","author":"Zhao","year":"1996","journal-title":"J. Biol. Chem."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"220","DOI":"10.1016\/S0014-5793(01)02980-5","article-title":"Green fluorescent protein as an indicator to monitor membrane protein overexpression in Escherichia coli","volume":"507","author":"Drew","year":"2001","journal-title":"FEBS Lett."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"13936","DOI":"10.1073\/pnas.0704546104","article-title":"High-throughput fluorescent-based optimization of eukaryotic membrane protein overexpression and purification in Saccharomyces cerevisiae","volume":"104","author":"Newstead","year":"2007","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"673","DOI":"10.1016\/j.str.2006.01.013","article-title":"Fluorescence-detection size-exclusion chromatography for precrystallization screening of integral membrane proteins","volume":"14","author":"Kawate","year":"2006","journal-title":"Structure"},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Becares, E.R., Pedersen, P.A., Gourdon, P., and Gotfryd, K. (2021). Overproduction of Human Zip (SLC39) Zinc Transporters in Saccharomyces cerevisiae for Biophysical Characterization. Cells, 10.","DOI":"10.3390\/cells10020213"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1309","DOI":"10.1002\/pro.2507","article-title":"Method to increase the yield of eukaryotic membrane protein expression in Saccharomyces cerevisiae for structural and functional studies","volume":"23","author":"Parker","year":"2014","journal-title":"Protein Sci."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"784","DOI":"10.1038\/nprot.2008.44","article-title":"GFP-based optimization scheme for the overexpression and purification of eukaryotic membrane proteins in Saccharomyces cerevisiae","volume":"3","author":"Drew","year":"2008","journal-title":"Nat. Protoc."},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Vidali, L., Rounds, C.M., Hepler, P.K., and Bezanilla, M. (2009). Lifeact-mEGFP reveals a dynamic apical F-actin network in tip growing plant cells. PLoS ONE, 4.","DOI":"10.1371\/journal.pone.0005744"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"8670","DOI":"10.1128\/JVI.74.18.8670-8679.2000","article-title":"Localization of Human Immunodeficiency Virus Type 1 Gag and Env at the Plasma Membrane by Confocal Imaging","volume":"74","author":"Resh","year":"2000","journal-title":"J. Virol."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"4427","DOI":"10.1091\/mbc.e03-04-0221","article-title":"Visualization of Protein Compartmentation within the Plasma Membrane of Living Yeast Cells","volume":"14","author":"Tanner","year":"2003","journal-title":"Mol. Biol. Cell"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"5044","DOI":"10.1128\/MCB.17.9.5044","article-title":"Zap1p, a metalloregulatory protein involved in zinc-responsive transcriptional regulation in Saccharomyces cerevisiae","volume":"17","author":"Zhao","year":"1997","journal-title":"Mol. Cell. Biol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"2419","DOI":"10.1056\/NEJMoa1510093","article-title":"Adapted Treatment Guided by Interim PET-CT Scan in Advanced Hodgkin\u2019s Lymphoma","volume":"374","author":"Johnson","year":"2016","journal-title":"N. Engl. J. Med."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"24466","DOI":"10.1074\/jbc.M110.109801","article-title":"Identification and Characterization of a Novel Human PP1 Phosphatase Complex","volume":"285","author":"Lee","year":"2010","journal-title":"J. Biol. Chem."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"69","DOI":"10.1016\/j.envexpbot.2018.09.020","article-title":"NtZIP11, a new Zn transporter specifically upregulated in tobacco leaves by toxic Zn level","volume":"157","author":"Kozak","year":"2018","journal-title":"Environ. Exp. Bot."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"53","DOI":"10.1016\/j.fgb.2012.11.004","article-title":"OmZnT1 and OmFET, two metal transporters from the metal-tolerant strain Zn of the ericoid mycorrhizal fungus Oidiodendron maius, confer zinc tolerance in yeast","volume":"52","author":"Khouja","year":"2013","journal-title":"Fungal Genet. Biol."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"11979","DOI":"10.1038\/ncomms11979","article-title":"Structural insights of ZIP4 extracellular domain critical for optimal zinc transport","volume":"7","author":"Zhang","year":"2016","journal-title":"Nat. Commun."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"18","DOI":"10.1016\/j.neuropharm.2016.05.016","article-title":"Isolation of proflavine as a blocker of G protein-gated inward rectifier potassium channels by a cell growth-based screening system","volume":"109","author":"Kawada","year":"2016","journal-title":"Neuropharmacology"}],"container-title":["Biomolecules"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2218-273X\/12\/5\/726\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T23:16:01Z","timestamp":1760138161000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2218-273X\/12\/5\/726"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,5,21]]},"references-count":46,"journal-issue":{"issue":"5","published-online":{"date-parts":[[2022,5]]}},"alternative-id":["biom12050726"],"URL":"https:\/\/doi.org\/10.3390\/biom12050726","relation":{},"ISSN":["2218-273X"],"issn-type":[{"type":"electronic","value":"2218-273X"}],"subject":[],"published":{"date-parts":[[2022,5,21]]}}}