Protein Biochemistry and Expression Regulation of Cadmium/Zinc Pumping ATPases in the Hyperaccumulator Plants Arabidopsis halleri and Noccaea caerulescens

• Published in: Frontiers in plant science Vol. 8; p. 835
• Main Authors: Mishra, Seema, Mishra, Archana, Küpper, Hendrik
• Format: Journal Article
• Language: English
• Published: Switzerland Frontiers Media S.A 22.05.2017
• Subjects: cadmium; heavy metal ATPase; metal hyperaccumulator plants; Plant Science; QISH; RT-qPCR; zinc; QISH; metal hyperaccumulator plants; cadmium; RT-qPCR; zinc; heavy metal ATPase
• ISSN: 1664-462X; 1664-462X
• DOI: 10.3389/fpls.2017.00835

P -ATPases are decisive for metal accumulation phenotypes, but mechanisms of their regulation are only partially understood. Here, we studied the Cd/Zn transporting ATPases NcHMA3 and NcHMA4 from as well as AhHMA3 and AhHMA4 from . Protein biochemistry was analyzed on HMA4 purified from roots of in active state. Metal titration of NcHMA4 protein with an electrochromic dye as charge indicator suggested that HMA4 reaches maximal ATPase activity when all internal high-affinity Cd binding sites are occupied. Although HMA4 was reported to be mainly responsible for xylem loading of heavy metals for root to shoot transport, the current study revealed high expression of NcHMA4 in shoots as well. Further, there were additional 20 and 40 kD fragments at replete Zn and toxic Cd , but not at deficient Zn concentrations. Altogether, the protein level expression analysis suggested a more multifunctional role of NcHMA4 than previously assumed. Organ-level transcription analysis through quantitative PCR of mRNA in and confirmed the strong shoot expression of both and . Further, in shoots was more abundant in 10 μM Zn and in Zn deficiency. In roots, was up-regulated in response to deficient Zn when compared to replete Zn and toxic Cd treatment. In both species, was much more expressed in shoots than in roots, and transcript levels remained rather constant regardless of Zn supply, but were up-regulated by 10 μM Cd . Analysis of cellular expression by quantitative mRNA hybridisation showed that in , both and mRNA levels were highest in the mesophyll, while in they were highest in the bundle sheath of the vein. This is likely related to the different final storage sites for hyperaccumulated metals in both species: epidermis in , mesophyll in .