Poster: Membrane transport

Abs # 168: Functional role of HMA2, a Zn transporting P-type ATPase

Presenter:	Eren, Elif , elif@wpi.edu
Authors	Eren, Elif  (A)   Orofino, Maria J. (A)   Arguello, Jose M. (A)
Affiliations:	(A): Worcester Polytechnic Institute

P_IB-type ATPases transport various heavy metal ions (Cu⁺, Cu²⁺, Zn²⁺, Cd²⁺, Co²⁺, etc.) across biological membranes. Several members of this subfamily are present in plants. Higher plants appear to be the only eukaryotes where Zn-ATPases are present. We have cloned and characterized HMA2 from Arabidopsis thaliana. HMA2 was first heterologously expressed in yeast and studied in isolated membrane fractions. HMA2 is a Zn-ATPase that is also activated by Cd²⁺ and to a lesser extent by some other divalent metals (Pd²⁺, Ni²⁺, Cu²⁺). The enzyme forms stable phosphorylated intermediates and interacts with Zn²⁺ and Cd²⁺ with high affinity (Zn²⁺ K_1/2= 0.11±0.03 µM and Cd²⁺ K_1/2= 0.028±0.007 µM). Its turnover number, similar to other P_IB-ATPases, is 175 1/min. HMA2 activity is dependent on milimolar concentration of cysteine and it is inhibited by vanadate. Analysis of transcript levels by RT-PCR suggests that the enzyme is present in all plant organs at all development stages. HMA2 mRNA levels did not change in plants exposed to various metals. Removal of HMA2 full-length transcript resulted in severe impairment of Zn homeostasis and accumulation of Zn²⁺ particularly in roots. Altogether, these results suggest that HMA2 is a housekeeping enzyme responsible for Zn²⁺ efflux from the cells and therefore required for normal Zn²⁺ homeostasis.