@phdthesis{Hoffmann2007,
  author    = {Hoffmann, Toni},
  title     = {Cloning and characterisation of the HMA3 gene and its promoter from Arabidopsis halleri (L.) O'Kane and Al'Shehbaz and Arabidopsis thaliana (L.) Heynhold},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus-15259},
  school      = {Universit{\"a}t Potsdam},
  year      = {2007},
  abstract  = {Being living systems unable to adjust their location to changing environmental conditions, plants display homeostatic networks that have evolved to maintain transition metal levels in a very narrow concentration range in order to avoid either deficiency or toxicity. Hence, plants possess a broad repertoire of mechanisms for the cellular uptake, compartmentation and efflux, as well as for the chelation of transition metal ions. A small number of plants are hypertolerant to one or a few specific transition metals. Some metal tolerant plants are also able to hyperaccumulate metal ions. The Brassicaceae family member Arabidopis halleri ssp. halleri (L.) O´KANE and AL´SHEHBAZ is a hyperaccumulator of zinc (Zn), and it is closely related to the non-hypertolerant and non-hyperaccumulating model plant Arabidopsis thaliana (L.) HEYNHOLD. The close relationship renders A. halleri a promising emerging model plant for the comparative investigation of the molecular mechanisms behind hypertolerance and hyperaccumulation. Among several potential candidate genes that are probably involved in mediating the zinc-hypertolerant and zinc-hyperaccumulating trait is AhHMA3. The AhHMA3 gene is highly similar to AtHMA3 (AGI number: At4g30120) in A. thaliana, and its encoded protein belongs to the P-type IB ATPase family of integral membrane transporter proteins that transport transition metals. In contrast to the low AtHMA3 transcript levels in A. thaliana, the gene was found to be constitutively highly expressed across different Zn treatments in A. halleri, especially in shoots. In this study, the cloning and characterisation of the HMA3 gene and its promoter from Arabidopsis halleri (L.) O´KANE and AL´SHEHBAZ and Arabidopsis thaliana (L.) HEYNHOLD is described. Heterologously expressed AhHMA3 mediated enhanced tolerance to Zn and to a much lesser degree to cadmium (Cd) but not to cobalt (Co) in metal-sensitive mutant strains of budding yeast. It is demonstrated that the genome of A. halleri contains at least four copies of AhHMA3, AhHMA3-1 to AhHMA3-4. A copy-specific real-time RT-PCR indicated that an AhHMA3-1 related gene copy is the source of the constitutively high transcript level in A. halleri and not a gene copy similar to AhHMA3-2 or AhHMA3-4. In accordance with the enhanced AtHMA3mRNA transcript level in A. thaliana roots, an AtHMA3 promoter-GUS gene construct mediated GUS activity predominantly in the vascular tissues of roots and not in shoots. However, the observed AhHMA3-1 and AhHMA3-2 promoter-mediated GUS activity in A. thaliana or A. halleri plants did not reflect the constitutively high expression of AhHMA3 in shoots of A. halleri. It is suggested that other factors e. g. characteristic sequence inserts within the first intron of AhHMA3-1 might enable a constitutively high expression. Moreover, the unknown promoter of the AhHMA3-3 gene copy could be the source of the constitutively high AhHMA3 transcript levels in A. halleri. In that case, the AhHMA3-3 sequence is predicted to be highly homologous to AhHMA3-1. The lack of solid localisation data for the AhHMA3 protein prevents a clear functional assignment. The provided data suggest several possible functions of the AhHMA3 protein: Like AtHMA2 and AtHMA4 it might be localised to the plasma membrane and could contribute to the efficient translocation of Zn from root to shoot and/or to the cell-to-cell distribution of Zn in the shoot. If localised to the vacuolar membrane, then a role in maintaining a low cytoplasmic zinc concentration by vacuolar zinc sequestration is possible. In addition, AhHMA3 might be involved in the delivery of zinc ions to trichomes and mesophyll leaf cells that are major zinc storage sites in A. halleri.},
  language  = {en}
}