The Roles of ZnT1 and ZnT4 in Glucose-Stimulated Zinc Secretion in Prostate Epithelial Cells
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RESEARCH ARTICLE
The Roles of ZnT1 and ZnT4 in Glucose-Stimulated Zinc Secretion in Prostate Epithelial Cells Su-Tang Lo,1,2 Daniel Parrott,1,2 M. Veronica Clavijo Jordan,1,2,3 Diya Binoy Joseph,3 Douglas Strand,3 U-Ging Lo,4 Ho Lin,5 Anza Darehshouri,6 A. Dean Sherry 1,2,7 1
Advanced Imaging Research Center, The University of Texas Southwestern Medical Center, 5323 Harry Hines Blvd, NE 4.210, Dallas, TX, 75390-8568, USA 2 Department of Radiology, UT Southwestern Medical Center, Dallas, TX, 75390-8896, USA 3 Athinoula A. Martinos Center for Biomedical Imaging, Massachusetts General Hospital, Harvard Medical School, Charlestown, MA, 02129, USA 4 Department of Urology, UT Southwestern Medical Center, Dallas, TX, 75390-9110, USA 5 Department of Life Sciences, National Chung Hsing University, Taichung City, 402, Taiwan 6 Electron Microscopy Core Facility, UT Southwestern Medical Center, Dallas, TX, 75390-9039, USA 7 Department of Chemistry, University of Texas at Dallas, Richardson, TX, 75083, USA
Abstract Purpose: We have previously demonstrated by MRI that high glucose stimulates efflux of zinc ions from the prostate. To our knowledge, this phenomena had not been reported previously and the mechanism remains unknown. Here, we report some initial observations that provide new insights into zinc processing during glucose-stimulated zinc secretion (GSZS) in the immortalized human prostate epithelial cell line, PNT1A. Additionally, we identified the subtypes of zinccontaining cells in human benign prostatic hyperplasia (BPH) tissue to further identify which cell types are likely responsible for zinc release in vivo. Procedure: An intracellular fluorescence marker, FluoZin-1-AM, was used to assess the different roles of ZnT1 and ZnT4 in zinc homeostasis in wild type (WT) and mRNA knockdown PNT1A cell lines. Additionally, Bafilomycin A1 (Baf) was used to disrupt lysosomes and assess the role of lysosomal storage during GSZS. ZIMIR, an extracellular zinc-responsive fluorescent marker, was used to assess dynamic zinc efflux of WT and ZnT1 mRNA knockdown cells exposed to high glucose. Electron microscopy was used to assess intracellular zinc storage in response to high glucose and evaluate how Bafilomycin A1 affects zinc trafficking. BPH cells were harvested from transurtheral prostatectomy tissue and stained with fluorescent zinc granule indicator (ZIGIR), an intracellular zinc-responsive fluorescent marker, before being sorted for cell types using flow cytometry. Results: Fluorescent studies demonstrate that ZnT1 is the major zinc efflux transporter in prostate epithelial cells and that loss of ZnT1 via mRNA knockdown combined with lysosomal storage disruption results in a nearly 4-fold increase in cytosolic zinc. Knockdown of ZnT1 dramatically reduces zinc efflux during GSZS. Electron microscopy (EM) reveals that glucose
Supplementary Information The online version contains supplementary material available at https://doi.org/10.1007/s11307-020-01557-x. Correspondence to: A. Sherry; e-mail: [email protected]
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