Development of Zn-related necrosis in tobacco is enhanced by expressing AtHMA4 and depends on the apoplastic Zn levels

AtHMA4 was previously shown to contribute to the control of Zn root-to-shoot translocation and tolerance to high Zn. However, heterologous expression of 35S::AtHMA4 in tobacco (Nicotiana tabacum cv. Xanthi) results in enhanced Zn sensitivity. This study provides a better understanding of the development of this Zn-sensitive phenotype and demonstrates that substantial modifications of Zn homeostasis occur due to AtHMA4 expression. We show that ectopically expressing AtHMA4 in tobacco results in overloading the root and leaf apoplast with Zn. The tissue and cellular distribution of Zn, monitored using Zinpyr-1, was altered in the AtHMA4-expressing plants compared with wild type. Increased loading of the leaf apoplast with Zn in AtHMA4 transformants induced necrosis; this appeared at lower levels of Zn supply in the transgenics compared with wild type. This study suggests that Zn concentration may be sensed in the apoplast of leaves, and if concentrations are above a certain threshold then particular groups of cells accumulate Zn and necrosis is initiated. Therefore, this could be considered as a mechanism for protecting the other parts of the photosynthetically active leaf from Zn toxicity.