AUTHOR=Iqbal Mazhar , Nawaz Ismat , Hassan Zeshan , Hakvoort Henk W., Bliek Mattijs , Aarts Mark G., Schat Henk 

TITLE=Expression of HMA4 cDNAs of the zinc hyperaccumulator Noccaea caerulescens from endogenous NcHMA4 promoters does not complement the zinc-deficiency phenotype of the Arabidopsis thaliana hma2hma4 double mutant

JOURNAL=Frontiers in Plant Science

VOLUME=4

YEAR=2013

URL=https://www.frontiersin.org/journals/plant-science/articles/10.3389/fpls.2013.00404

DOI=10.3389/fpls.2013.00404

ISSN=1664-462X

ABSTRACT=<p><italic>Noccaea caerulescens</italic> (<italic>Nc</italic>) exhibits a very high constitutive expression of the heavy metal transporting ATPase, <italic>HMA4</italic>, as compared to the non-hyperaccumulator <italic>Arabidopsis thaliana</italic> (<italic>At</italic>), due to copy number expansion and altered <italic>cis</italic>-regulation. We screened a BAC library for <italic>HMA4</italic> and found that <italic>HMA4</italic> is triplicated in the genome of a <italic>N. caerulescens</italic> accession from a former Zn mine near La Calamine (LC), Belgium. We amplified multiple <italic>HMA4</italic> promoter sequences from three calamine <italic>N. caerulescens</italic> accessions, and expressed <italic>AtHMA4</italic> and different <italic>NcHMA4</italic> cDNAs under <italic>At</italic> and <italic>Nc</italic><italic>HMA4</italic> promoters in the <italic>A. thaliana</italic> (Col) <italic>hma2hma4</italic> double mutant. Transgenic lines expressing <italic>HMA4</italic> under the <italic>At</italic> promoter were always fully complemented for root-to-shoot Zn translocation and developed normally at a 2-μM Zn supply, whereas the lines expressing <italic>HMA4</italic> under <italic>Nc</italic> promoters usually showed only slightly enhanced root to shoot Zn translocation rates in comparison with the double mutant, probably owing to ectopic expression in the roots, respectively. When expression of the Zn deficiency responsive marker gene <italic>ZIP4</italic> was tested, the transgenic lines expressing <italic>AtHMA4</italic> under an <italic>NcHMA4-1-LC</italic> promoter showed on average a 7-fold higher expression in the leaves, in comparison with the double <italic>hma2hma4</italic> mutant, showing that this construct aggravated, rather than alleviated the severity of foliar Zn deficiency in the mutant, possible owing to expression in the leaf mesophyll.</p>