ZIP3 is essential for the encephalitogenic function of pTh17 cells by regulating intracellular zinc levels

Zinc (Zn²⁺) is the second most abundant trace metal in eukaryotes and essential for many cellular functions. It possesses well-established immunomodulatory properties and disruption of Zn²⁺ signaling compromises immune functions resulting in immunodeficiency. In addition, Zn²⁺ has been implicated in several autoimmune diseases such as multiple sclerosis (MS).

We used a forward genomic screen to identify Zn²⁺ transporters that control the function of encephalitogenic T helper (Th) 17 cells, and thus, drive CNS inflammation in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS. We measured cytosolic Zn²⁺ levels in T cells with a genetically encoded Zn²⁺ reporter and used confocal microscopy to determine the subcellular localization of ZIP3. We used shRNA and CRISPR gene editing to delete Slc39a3 (encoding ZIP3) to determine its role in CD4⁺ T cells in vitro and pathogenic Th17 (pTh17) cells in vivo in EAE.

Using a shRNA screen in myelin-specific pTh17 cells in vivo, we found that ZIP3 is required for their expansion in the CNS of mice with EAE. When ectopically expressed, ZIP3 was located predominantly in the Golgi apparatus. Deletion of Slc39a3 resulted in decreased cytosolic Zn²⁺ levels selectively in pTh17 cells without affecting those in non-polarized CD4⁺ T cells. Slc39a3-deficient pTh17 cells showed an increased susceptibility to apoptotic cell death and reduced CD69 expression in vitro, while proliferation and cytokine production remained intact. Slc39a3 deletion in pTh17 cells significantly reduced the severity of EAE by reducing leukocyte infiltration of the CNS.

ZIP3 is essential for the encephalitogenic function of pTh17 cells and promoting EAE, likely by regulating cytosolic Zn²⁺ levels.