IL-4 Induces Metallothionein 3- and SLC30A4-Dependent Increase in Intracellular Zn2+ that Promotes Pathogen Persistence in Macrophages

SummaryAlternative activation of macrophages promotes wound healing but weakens antimicrobial defenses against intracellular pathogens. The mechanisms that suppress macrophage function to create a favorable environment for pathogen growth remain elusive. We show that interleukin (IL)-4 triggers a metallothionein 3 (MT3)- and Zn exporter SLC30A4-dependent increase in the labile Zn2+ stores in macrophages and that intracellular pathogens can exploit this increase in Zn to survive. IL-4 regulates this pathway by shuttling extracellular Zn into macrophages and by activating cathepsins that act on MT3 to release bound Zn. We show that IL-4 can modulate Zn homeostasis in both human monocytes and mice. In vivo, MT3 can repress macrophage function in an M2-polarizing environment to promote pathogen persistence. Thus, MT3 and SLC30A4 dictate the size of the labile Zn2+ pool and promote the survival of a prototypical intracellular pathogen in M2 macrophages

Metallothioneins: Emerging Modulators in Immunity and Infection

Metallothioneins (MTs) are a family of metal-binding proteins virtually expressed in all organisms including prokaryotes, lower eukaryotes, invertebrates and mammals. These proteins regulate homeostasis of zinc (Zn) and copper (Cu), mitigate heavy metal poisoning, and alleviate superoxide stress. In recent years, MTs have emerged as an important, yet largely underappreciated, component of the immune system. Innate and adaptive immune cells regulate MTs in response to stress stimuli, cytokine signals and microbial challenge. Modulation of MTs in these cells in turn regulates metal ion release, transport and distribution, cellular redox status, enzyme function and cell signaling. While it is well established that the host strictly regulates availability of metal ions during microbial pathogenesis, we are only recently beginning to unravel the interplay between metal-regulatory pathways and immunological defenses. In this perspective, investigation of mechanisms that leverage the potential of MTs to orchestrate inflammatory responses and antimicrobial defenses has gained momentum. The purpose of this review, therefore, is to illumine the role of MTs in immune regulation. We discuss the mechanisms of MT induction and signaling in immune cells and explore the therapeutic potential of the MT-Zn axis in bolstering immune defenses against pathogens

Schematic of Zn regulation in phagocytes.

<p>Mechanisms of Zn regulation in phagocytes, grouped into three categories: Zn²⁺ transport, storage, and binding. (<b>A</b>) Zn²⁺ transport across the cell membrane is mediated by ZIPs and ZNTs. (<b>B</b>) Intracellular Zn²⁺ is transported into and stored in organelles such as endosomes, lysosomes, Golgi, and zincosomes by various transporters represented in the figure; the transporters that mediate Zn²⁺ flux across zincosomes have not been identified. (<b>C</b>) Zn²⁺ is bound and sequestered by intracellular or secreted metal binding proteins such as MTs and calprotectin.</p

Schematic of Zn regulation in activated macrophages infected with a fungal pathogen.

<p>Zn regulation in a GM-CSF–activated macrophage leading to defense against fungal infection. (<b>A</b>) GM-CSF binds to the GM-CSF receptor on infected macrophages, activates STAT3 and STAT5 signaling, and triggers transcriptional activation in the nucleus. (<b>B</b>) Induction of ZIP2 causes increased Zn²⁺ influx, which may support increased metabolic functions to cope with stress in the infected macrophage. (<b>C</b>) STAT3 and STAT5 induce expression of MTs that sequester labile intracellular Zn²⁺. (<b>D</b>) Zn²⁺ is mobilized into the Golgi apparatus, associated with increased expression of Golgi membrane transporters ZNT4 and ZNT7. (<b>E</b>) Speculated lysosomal Zn deprivation by influx into the cytosol by ZIPs; the dotted arrow represents predicted sequestration of Zn²⁺ from this source by MTs. (<b>F</b>) Zn²⁺ inhibits proton flux via HV1, but the “Zn²⁺-deprived” environment lifts the inhibitory action (shown on extreme right of the phagolysosomal membrane) and H⁺ generated by Nox activity is channeled into phagolysosomes effectively sustaining production of superoxide radicals by the enzyme. (<b>G</b>) The pathogen senses a Zn²⁺-deprived environment and activates Zn-responsive transcription machinery to trigger Zn²⁺ import via fungal transporters and zincophore systems; ultimately, deficiency of Zn²⁺ starves the pathogen of this metal and simultaneously enhances superoxide burst in phagocytes, culminating into inhibition of fungal growth.</p