Metalation and activation of Zn²⁺enzymes via early secretory pathway-resident ZNT proteins

Zinc (Zn²⁺), an essential trace element, binds to various proteins, including enzymes, transcription factors, channels, and signaling molecules and their receptors, to regulate their activities in a wide range of physiological functions. Zn²⁺ proteome analyses have indicated that approximately 10% of the proteins encoded by the human genome have potential Zn²⁺ binding sites. Zn²⁺binding to the functional site of a protein (for enzymes, the active site) is termed Zn²⁺metalation. In eukaryotic cells, approximately one-third of proteins are targeted to the endoplasmic reticulum; therefore, a considerable number of proteins mature by Zn²⁺metalation in the early secretory pathway compartments. Failure to capture Zn²⁺ in these compartments results in not only the inactivation of enzymes (apo-Zn²⁺ enzymes), but also their elimination via degradation. This process deserves attention because many Zn²⁺ enzymes that mature during the secretory process are associated with disease pathogenesis. However, how Zn²⁺is mobilized via Zn²⁺ transporters, particularly ZNTs, and incorporated in enzymes has not been fully elucidated from the cellular perspective and much less from the biophysical perspective. This review focuses on Zn²⁺ enzymes that are activated by Zn²⁺ metalation via Zn²⁺ transporters during the secretory process. Further, we describe the importance of Zn²⁺ metalation from the physiopathological perspective, helping to reveal the importance of understanding Zn²⁺ enzymes from a biophysical perspective

Zinc transport via ZNT5-6 and ZNT7 is critical for cell surface glycosylphosphatidylinositol-anchored protein expression

Glycosylphosphatidylinositol (GPI)-anchored proteins play crucial roles in various enzyme activities, cell signaling and adhesion, and immune responses. While the molecular mechanism underlying GPI-anchored protein biosynthesis has been well studied, the role of zinc transport in this process has not yet been elucidated. Zn transporter (ZNT) proteins mobilize cytosolic zinc to the extracellular space and to intracellular compartments. Here, we report that the early secretory pathway ZNTs [ZNT5-ZNT6 heterodimers (ZNT5-6) and ZNT7-ZNT7 homodimers (ZNT7)], which supply zinc to the lumen of the early secretory pathway compartments are essential for GPI-anchored protein expression on the cell surface. We show, using overexpression and gene disruption/re-expression strategies in cultured human cells, that loss of ZNT5-6 and ZNT7 zinc transport functions results in significant reduction in GPI-anchored protein levels similar to that in mutant cells lacking phosphatidylinositol glycan anchor biosynthesis (PIG) genes. Furthermore, medaka fish with disrupted Znt5 and Znt7 genes show touch-insensitive phenotypes similar to zebrafish Pig mutants. These findings provide a previously unappreciated insight into the regulation of GPI-anchored protein expression and protein quality control in the early secretory pathway

Pigmentation and TYRP1 expression are mediated by zinc through the early secretory pathway-resident ZNT proteins

メラニン生合成には、亜鉛も必要だった！. 京都大学プレスリリース. 2023-04-19.Tyrosinase (TYR) and tyrosinase-related proteins 1 and 2 (TYRP1 and TYRP2) are essential for pigmentation. They are generally classified as type-3 copper proteins, with binuclear copper active sites. Although there is experimental evidence for a copper cofactor in TYR, delivered via the copper transporter, ATP7A, the presence of copper in TYRP1 and TYRP2 has not been demonstrated. Here, we report that the expression and function of TYRP1 requires zinc, mediated by ZNT5–ZNT6 heterodimers (ZNT5–6) or ZNT7–ZNT7 homodimers (ZNT7). Loss of ZNT5–6 and ZNT7 function results in hypopigmentation in medaka fish and human melanoma cells, and is accompanied by immature melanosomes and reduced melanin content, as observed in TYRP1 dysfunction. The requirement of ZNT5–6 and ZNT7 for TYRP1 expression is conserved in human, mouse, and chicken orthologs. Our results provide novel insights into the pigmentation process and address questions regarding metalation in tyrosinase protein family

Metalation and activation of Zn²⁺enzymes via early secretory pathway-resident ZNT proteins

Zinc (Zn²⁺), an essential trace element, binds to various proteins, including enzymes, transcription factors, channels, and signaling molecules and their receptors, to regulate their activities in a wide range of physiological functions. Zn²⁺ proteome analyses have indicated that approximately 10% of the proteins encoded by the human genome have potential Zn²⁺ binding sites. Zn²⁺binding to the functional site of a protein (for enzymes, the active site) is termed Zn²⁺metalation. In eukaryotic cells, approximately one-third of proteins are targeted to the endoplasmic reticulum; therefore, a considerable number of proteins mature by Zn²⁺metalation in the early secretory pathway compartments. Failure to capture Zn²⁺ in these compartments results in not only the inactivation of enzymes (apo-Zn²⁺ enzymes), but also their elimination via degradation. This process deserves attention because many Zn²⁺ enzymes that mature during the secretory process are associated with disease pathogenesis. However, how Zn²⁺is mobilized via Zn²⁺ transporters, particularly ZNTs, and incorporated in enzymes has not been fully elucidated from the cellular perspective and much less from the biophysical perspective. This review focuses on Zn²⁺ enzymes that are activated by Zn²⁺ metalation via Zn²⁺ transporters during the secretory process. Further, we describe the importance of Zn²⁺ metalation from the physiopathological perspective, helping to reveal the importance of understanding Zn²⁺ enzymes from a biophysical perspective

Patients with CDH23

Conclusions: CDH23 mutations and the 1555A>G mitochondrial mutation were identified among our series of electric acoustic stimulation (EAS) patients, confirming that these genes were important in hearing loss with involvement of high frequency. Successful hearing preservation as well as good outcomes from EAS indicated that patients with this combination of mutations are good candidates for EAS. Objectives: Screening for gene mutations that possibly cause hearing loss involving high frequency was performed to identify the responsible genes in patients with EAS. In addition to a review of the genetic background of the patients with residual hearing loss, the benefit of EAS for patients with particular gene mutations was evaluated. Methods: Eighteen patients (15 late-onset, 3 early-onset) with residual hearing who had received EAS were included in this study. Genetic analysis was performed to identify GJB2, CDH23, SLC26A4, and the 1555 mitochondrial mutations. Results: Three early-onset patients had CDH23 mutations. One late-onset patient had the 1555 A>G mitochondrial mutation