Unravelling zinc homeostatic mechanisms in the crop plant barley

Abstract

Zinc (Zn) is an essential micronutrient in plants but becomes toxic when present in excess, with nutritional extremes leading to agricultural yield losses. Homeostatic mechanisms are in place to control cellular Zn levels with transcription factors and membrane transport proteins playing vital roles. In Arabidopsis thaliana, two F-group bZIP transcription factors, bZIP19 and bZIP23, are proposed to sense and respond to Zn deficiency by regulating the expression of particular Zn membrane transporters, ZIPs (ZRT/IRT-like proteins). In this thesis, four unique bzip19 bzip23 knockout mutants with different combinations of T-DNA insertion sites were generated and shown to be hypersensitive to Zn-deficiency. To understand the role of F-group bZIPs in the economically important crop Hordeum vulgare (barley), HvbZIP1, 10, 56, 57, 58, and 61 were cloned and characterized to various extents. HvbZIP56, HvbZIP57, HvbZIP62 but not HvbZIP1 partially rescue the hypersensitive phenotype of the A. thaliana bzip19-4 bzip23-2 mutant. HvbZIP56 was localised to the cytoplasm and nucleus when expressed in A. thaliana and tobacco. Promoter analysis demonstrates that barley ZIP transporters that are up-regulated under Zn deficiency contain cis Zn-deficiency response elements (ZDREs), similarly to A. thaliana. Overall, these results indicate that the mechanisms operating in controlling Zn levels in barley are conserved.

Two transporters, AtHMA3 (a P1B-2-type ATPase) and AtMTP1 (a Metal tolerance protein) have been implicated in sequestering Zn in Arabidopsis vacuoles to alleviate Zn toxicity. In this study, only the mtp1 mutant and not the hma3 mutant showed hypersensitivity to high Zn levels. HvHMA3 (a barley P1B-2-ATPase) when expressed in mtp1 rescued this hypersensitivity indicating a role in Zn transport, although this could not be confirmed by expression in S. cerevisiae. This study represents a significant step forward in understanding the mechanisms controlling Zn responses in cereal crops, and will aid in developing strategies for crop improvement.

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