Poplar interactions with zinc for use in bioremediation and monitoring

Plant mechanisms regulating environmental heavy metal interactions are vital for plant survival. Plants must maintain adequate metal levels while preventing excesses. Several mechanisms involved in heavy metal uptake and sequestration have been identified and studied in hyperaccumulating plants such as Thlaspi caerulescens. These plants accumulate large quantities of metals, but their use in remediation is limited by their small size. On the other hand, mechanisms in high-biomass, non-hyperaccumulating perennial species such as poplar (Populus spp.) are unknown. The central goal of this project is to delineate specific mechanisms in poplar regulating the heavy metal zinc (Zn) for potential use in bioremediation and real-time monitoring. Specifically, project aims are: 1) Determine the role of HMA4 and PCS1 genes in poplar; 2) Delineate the ZIP gene family including ZIP1 and ZNT1 activity; and 3) Harness fluorescent energy transfer to engineer a poplar tree that monitors Zn-soil contamination. These are addressed using current technologies including phylogenetic analysis, gene transformation, expression assays, promoter-GUS assays, fluorescent-gene imaging, and metal assays. Through these experiments, mechanisms controlling heavy metal interaction are identified and characterized in poplar. Poplar contains a large number of genes in both the ZIP and HMA4 families, but only two members in the PCS family. Poplar also contains several genes that share close sequence and structural homology to those in hyperaccumulators. However, there is an overall divergence from hyperaccumulators in regards to expression across an environmental Zn gradient. Poplar tightly regulates Zn intake by suppressing absorption avenues under Zn excess. Over-expression of HMA4 and PCS1 resulted in more tolerance and more accumulation, respectively, in poplar lines. These findings support a regulatory system used in poplar to limit Zn under excess and promote Zn under deficiency. Using ZNT1 and its natural expression gradient, a chimeric protein was created that served as a biosensor in both poplar and Arabidopsis thaliana host plants and was able to discriminate between 1μM and 10mM Zn concentrations. These findings add to current knowledge of heavy metal regulation and help fill the gap of knowledge currently existing on the regulatory mechanisms that perennial trees use to control heavy metals