ZIP11 Regulates Nuclear Zinc Homeostasis in HeLa Cells and Is Required for Proliferation and Establishment of the Carcinogenic Phenotype

Abstract

Zinc (Zn) is an essential trace element that plays a key role in several biological processes, including transcription, signaling, and catalysis. A subcellular network of transporters ensures adequate distribution of Zn to facilitate homeostasis. Among these are a family of importers, the Zrt/Irt-like proteins (ZIP), which consists of 14 members (ZIP1-ZIP14) that mobilize Zn from the extracellular domain and organelles into the cytosol. Expression of these transporters varies among tissues and during developmental stages, and their distribution at various cellular locations is essential for defining the net cellular Zn transport. Normally, the ion is bound to proteins or sequestered in organelles and vesicles. However, though research has focused on Zn internalization in mammalian cells, little is known about Zn mobilization within organelles, including within the nuclei under both normal and pathological conditions. Analyses from stomach and colon tissues isolated from mouse suggested that ZIP11 is the only ZIP transporter localized to the nucleus of mammalian cells, yet no clear cellular role has been attributed to this protein. We hypothesized that ZIP11 is essential to maintaining nuclear Zn homeostasis in mammalian cells. To test this, we utilized HeLa cells, as research in humans correlated elevated expression of ZIP11 with poor prognosis in cervical cancer patients. We stably knocked down ZIP11 in HeLa cancer cells and investigated the effect of Zn dysregulation in vitro. Our data show that ZIP11 knockdown (KD) reduced HeLa cells proliferation due to nuclear accumulation of Zn. RNA-seq analyses revealed that genes related to angiogenesis, apoptosis, mRNA metabolism, and signaling pathways are dysregulated. Although the KD cells undergoing nuclear Zn stress can activate the homeostasis response by MTF1 and MT1, the RNA-seq analyses showed that only ZIP14 (an importer expressed on the plasma membrane and endocytic vesicles) is mildly induced, which may explain the sensitivity to elevated levels of extracellular Zn. Consequently, ZIP11 KD HeLa cells have impaired migration, invasive properties and decreased mitochondrial potential. Furthermore, KD of ZIP11 delayed cell cycle progression and rendered an enhanced senescent state in HeLa cells, pointing to a novel mechanism whereby maintenance of nuclear Zn homeostasis is essential for cancer progression.