Transcriptional activation and localization of expression of Brassica juncea putative metal transport protein BjMTP1.
BACKGROUND
Metal hyperaccumulators, including various Thlaspi species, constitutively express the putative metal transporter MTP1 to high levels in shoots. Here we present data on the transcriptional regulation and localization of expression of the homologous gene BjMTP1 in Brassica juncea. Though B. juncea lacks the ability to hyperaccumulate metals, its relatively high biomass, rapid growth and relatedness to true metal hyperaccumulating plants makes it a promising starting point for the development of plants for phytoremediation. Our goal in this study is to determine the transcriptional regulation of MTP1 in order to start to better understanding the physiological role of MTP1 in B. juncea.


RESULTS
Steady-state mRNA levels of BjMTP1 were found to be enhanced 8.8, 5.9, and 1.6-fold in five-day-old B. juncea seedlings after exposure to Ni2+, Cd2+ or Zn2+, respectively. This was also reflected in enhanced GUS activity in B. juncea seedlings transformed with BjMTP1 promoter::GUSPlus after exposure to these metals over a similar range of toxicities from mild to severe. However, no increase in GUS activity was observed after exposure of seedlings to cold or heat stress, NaCl or hydrogen peroxide. GUS expression in Ni2+ treated seedlings was localized in roots, particularly in the root-shoot transition zone. In four- week- old transgenic plants BjMTP1 promoter activity also primarily increased in roots in response to Ni2+ or Cd2+ in plants transformed with either GUS or mRFP1 as reporter genes, and expression was localized to the secondary xylem parenchyma. In leaves, BjMTP1 promoter activity in response to Ni2+ or Cd2+ spiked after 24 h then decreased. In shoots GUS expression was prominently present in the vasculature of leaves, and floral parts.


CONCLUSIONS
Our studies establish that a 983 bp DNA fragment upstream of the BjMTP1 translational start site is sufficient for the specific activation by Ni2+ and Cd2+ of BjMTP1 expression primarily in roots. Activation of expression by both metals in roots is primarily localized to the xylem parenchyma cells. This study is the first to identify specific Ni2+ and Cd2+ transcriptional regulation and tissue localization of BjMTP1.
Muthukumar
Balasubramaniam
B
Department of Horticulture and Landscape Architecture, Purdue University, West Lafayette, IN 47907-1392, USA. muthukum@purdue.edu
Yakubov
Bakhtiyor
B
Salt
David E
DE
D001483:Base Sequence; D001937:Brassica / Q000235:genetics*; D027682:Cation Transport Proteins / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers; D008670:Metals / Q000378:metabolism; D018517:Plant Roots / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D015533:Transcriptional Activation
D027682:Cation Transport Proteins; D017931:DNA Primers; D008670:Metals; D012333:RNA, Messenger