Characterization of ZIP genes responsible for zinc homeostasis in C. elegans

The essential element zinc plays an important structural and functional role in proteins in all living organisms. Zinc homeostasis is critical, because both zinc deficiency and excess are deleterious; in humans, defective zinc homeostasis leads to several disease states. The mechanisms utilized by animals to respond to excess zinc have been extensively characterized, but much less is known about how animals sense and respond to zinc deficiency. A primary method animals use to increase zinc content is through the zinc transport family known as the Zrt, Irt-like proteins (ZIPs). Caenorhabditis elegans is a powerful experimental system to study the mechanisms of zinc deficiency based on sophisticated genetic and cell biological approaches, and studies of C. elegans are likely to be relevant to humans, since both worms and humans have 14 ZIP family members. To characterize the mechanisms that animals use to respond to zinc deficiency, we examined the transcriptional response of the ZIP family members during zinc deficient conditions. We demonstrated that three ZIP genes in C. elegans, zipt-2.1, zipt-2.3, and zipt-7.1, are upregulated in zinc deficient conditions. The promoters of these genes contained a conservedcis-regulatory element we have named the low zinc activating (LZA) element. This element was necessary and sufficient to drive transcriptional activation in zinc deficient conditions. We also bioinformatically identified candidate genes that contained an LZA within their promoters and demonstrated that these genes are also activated by zinc deficient conditions. To understand the conservation of the function of the LZA, we transfected the promoter of zipt-2.3 into human cells and determined that the promoter was activated in zinc deficient conditions and this activation was dependent on the LZA element. These efforts elucidated the mechanisms that animals use to respond to zinc deficiency, including the discovery of a novel, conserved cis-regulatory element. Another mechanism used by animals to respond to changes in zinc availability is through the function of zinc transport proteins. We identified the C. elegans ZIP family member zipt-2.3 as a gene that was essential for growth and development during zinc deficiency. ZIPT-2.3 was expressed within lysosome-related organelles known as gut granules within the C. elegans intestine. This ZIP mediates the mobilization of zinc from these storage sites to allow animals to maintain proper zinc homeostasis. These results demonstrated that zinc storage and its subsequent mobilization from intracellular storage sites are the major mechanisms these animals use to adapt to changes in zinc status, and these mechanisms have been suggested to play crucial roles in other organisms