Background: Root-to-shoot translocation of zinc (Zn) and cadmium (Cd) depends on the concentrations of both metals in the medium. A previous study on tobacco (Nicotiana tabacum) pointed to the contribution of NtZIP1, NtZIP2, NtZIP4 and NtIRT1-like in the regulation of this phenomenon. To learn more, Zn and Cd accumulation, root/shoot distribution and the expression of ZIP genes were investigated in the apical, middle and basal root parts. Results: We show that Zn/Cd status-dependent root-shoot distribution of both metals was related to distinct metal accumulation in root parts. At low Zn and Cd in the medium, the apical part contained the highest metal level; at higher concentrations, the middle and basal parts were the major sink for excess metal. The above were accompanied by root part-specific expression pattern modifications of ZIPs (NtZIP1-like, NtZIP2, NtZIP4A/B, NtZIP5A/B, NtZIP5-like, NtZIP8, NtZIP11, NtIRT1, and NtIRT1-like) that fell into four categories with respect to the root part. Furthermore, for lower Zn/Cd concentrations changes were noted for NtZIP5A/B and NtZIP5-like only, but at higher Zn and Cd levels for NtZIP1-like, NtZIP5-like, NtZIP8, NtZIP11, NtIRT1, and NtIRT1-like. NtZIP1, here renamed to NtZIP5B, was cloned and characterized. We found that it was a zinc deficiency-inducible transporter involved in zinc and cadmium uptake from the soil solution primarily by the middle root part. Conclusions: We conclude that regulation of the longitudinal distribution of Zn and Cd is highly specific, and that the apical, middle and basal root parts play distinct roles in Zn/Cd status-dependent control of metal translocation efficiency to shoots, including the stimulation of Zn translocation to shoots in the presence of Cd. These results provide new insight into the root part-specific unique role of NtZIP5B and other ZIP genes in the longitudinal distribution of zinc and cadmium and their contribution to the regulation of root-to-shoot translocation.

In tobacco, the efficiency of Zn translocation to shoots depends on Zn/Cd status. Previous studies pointed to the specific contribution of root parts in the regulation of this process, as well as the role of NtZIP4A/B (from the ZIP family; Zrt Irt-like Proteins). Here, to verify this hypothesis, NtZIP4A/B RNAi lines were generated. Then, in plants exposed to combinations of Zn and Cd concentrations in the medium, the consequences of NtZIP4A/B suppression for the translocation of both metals were determined. Furthermore, the apical, middle, and basal root parts were examined for accumulation of both metals, for Zn localization (using Zinpyr-1), and for modifications of the expression pattern of ZIP genes. Our results confirmed the role of NtZIP4A/B in the control of Zn/Cd-status-dependent transfer of both metals to shoots. Furthermore, they indicated that the middle and basal root parts contributed to the regulation of this process by acting as a reservoir for excess Zn and Cd. Expression studies identified several candidate ZIP genes that interact with NtZIP4A/B in the root in regulating Zn and Cd translocation to the shoot, primarily NtZIP1-like in the basal root part and NtZIP2 in the middle one.

Metal tolerance proteins (MTPs) from the CDF (Cation Diffusion Facilitator) family are efflux transporters that play a crucial role in metal homeostasis by maintaining optimal metal concentrations in the cytoplasm. Here, a novel tobacco NtMTP2 transporter was cloned and characterized. It encodes a 512 aa protein containing all specific CDF family domains. A GFP-NtMTP2 fusion protein localizes to the tonoplast in tobacco cells. NtMTP2 expression in yeast conferred tolerance to Co and Ni, indicating that the protein mediates transport of both metals, but not Zn, Mn, Cu, Fe, or Cd. Nonetheless, the expression level was not affected by Co or Ni, except for an increase in leaves at high Co concentrations. Its expression in plant parts remained stable during development, but increased in the leaves of older plants. Analysis of tobacco expressing a promoter-GUS construct indicates that the main sites of promoter activity are the conductive tissue throughout the plant and the palisade parenchyma in leaves. Our results suggest that NtMTP2 is a tonoplast transporter mediating sequestration of Co and Ni into vacuoles and an important housekeeping protein that controls the basal availability of micronutrients and plays a role in the sequestration of metal excess, specifically in leaves.