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Title: Cooperative Functions of ZnT1, Metallothionein and ZnT4 in the Cytoplasm Are Required for Full Activation of TNAP in the Early Secretory Pathway
Authors: Fujimoto, Shigeyuki
Itsumura, Naoya
Tsuji, Tokuji
Anan, Yasumi
Tsuji, Natsuko
Ogra, Yasumitsu
Kimura, Tomoki
Miyamae, Yusaku
Masuda, Seiji  kyouindb  KAKEN_id
Nagao, Masaya  kyouindb  KAKEN_id
Kambe, Taiho  kyouindb  KAKEN_id
Author's alias: 神戸, 大朋
Issue Date: 18-Oct-2013
Publisher: Public Library of Science
Journal title: PLoS ONE
Volume: 8
Issue: 10
Thesis number: e77445
Abstract: The activation process of secretory or membrane-bound zinc enzymes is thought to be a highly coordinated process involving zinc transport, trafficking, transfer and coordination. We have previously shown that secretory and membrane-bound zinc enzymes are activated in the early secretory pathway (ESP) via zinc-loading by the zinc transporter 5 (ZnT5)-ZnT6 hetero-complex and ZnT7 homo-complex (zinc transport complexes). However, how other proteins conducting zinc metabolism affect the activation of these enzymes remains unknown. Here, we investigated this issue by disruption and re-expression of genes known to be involved in cytoplasmic zinc metabolism, using a zinc enzyme, tissue non-specific alkaline phosphatase (TNAP), as a reporter. We found that TNAP activity was significantly reduced in cells deficient in ZnT1, Metallothionein (MT) and ZnT4 genes (ZnT1^[−/−]MT^[−/−]ZnT4^[−/−] cells), in spite of increased cytosolic zinc levels. The reduced TNAP activity in ZnT1^[−/−]MT^[−/−]ZnT4^[−/−] cells was not restored when cytosolic zinc levels were normalized to levels comparable with those of wild-type cells, but was reversely restored by extreme zinc supplementation via zinc-loading by the zinc transport complexes. Moreover, the reduced TNAP activity was adequately restored by re-expression of mammalian counterparts of ZnT1, MT and ZnT4, but not by zinc transport-incompetent mutants of ZnT1 and ZnT4. In ZnT1^[−/−]MT^[−/−]ZnT4^[−/−] cells, the secretory pathway normally operates. These findings suggest that cooperative zinc handling of ZnT1, MT and ZnT4 in the cytoplasm is required for full activation of TNAP in the ESP, and present clear evidence that the activation process of zinc enzymes is elaborately controlled.
Rights: © 2013 Fujimoto et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
DOI(Published Version): 10.1371/journal.pone.0077445
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