Zip14 ablation modulates HDAC activity through zinc transport in intestinal epithelial cells
Authors: Jimenez-Rondan Felix , Ruggiero Courtney , Cousins Robert 
Published: May 2023

Zinc is known for its modulation of proinflammatory responses, in part by targeting nuclear factor kappa B (NF-ĸβ), signal transducer and activator of transcription 3 (STAT3) and expression of specific cytokines, including IFNγ. However, the molecular mechanism underlying Zn-induced cellular response to inflammation remains largely unknown. Our studies, using isolated mouse intestinal epithelial cells (IECs) and mouse intestinal organoids, both derived from Zip14 (Slc39a14) knockout mice, as well as intestinal-specific Zip14 knockout mice, with corresponding control genotypes, were used as models to evaluate the molecular basis for zinc-related inflammatory response in IECs, particularly a potential role for epigenetic modifications. Using chromatin immunoprecipitation (ChIP) assays employing chromatin from IECs, NF-ĸβ was found a major regulator of the Zip14 gene, at steady state. Compared to transcription factors (TFs) STAT3, ATF4 and ATF6, the involvement of NF-ĸβ supports the responsiveness of Zip14 expression with proinflammatory conditions. Similarly, more NF-ĸβ binding was found for iNOS, Cldn-2 and Cldn-8 promoters; all are genes that upregulate in IECs with ablation of Zip14. In contrast, markedly less NF-ĸβ was bound to the promoter of the defense gene IL-18 and C ldn-1; both of which were downregulated with Zip14 ablation. Differential gene expression with Zip14 ablation, as measured by RNA-seq, was limited to ~ 1% of total gene targets, at a false discovery rate of ± 2. RNA from control and Zip14 knockout IECs, intestinal organoids derived from those mice, and an IEC cell line treated with siRNA for Zip14, confirmed the differential expression profiles via qPCR. IECs from the models used here consistently show reduced histone deacetylase (HDAC) activity when Zip14 expression is limited. Moreover, that reduction in activity is concurrent with reduced zinc uptake as measured fluorometrically with the zinc probe, FluoZin-3. Additionally, histone acetylation is greater in IECs when Zip14 is deleted. Treatment of IECs with HDAC inhibitors support a link between Zip14 ablation and HDAC activity. Many HDACs are zinc-metalloenzymes and activity is inhibited with zinc chelation. We propose that deletion of Zip14 lessens HDAC activity, allowing for greater TF occupancy for promoters of specific genes. In summary, we demonstrate that histone acetylation is involved in a zinc-induced anti-inflammatory response in intestinal epithelial cells that is regulated by HDAC activity. Our study shows an important connection between histone modification and the pathological process of zinc-related intestinal inflammation and provides a mechanism for zinc-mediated epigenetic regulation in IECs. Supported by NIH grant DK 094244 and Boston Family Endowment Funds of the University of Florida Foundation. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.