Cucumber metal tolerance protein CsMTP9 is a plasma membrane H+-coupled antiporter involved in the Mn2+ and Cd2+ efflux from root cells

Abstract

Members of the plant metal tolerance protein (MTP) family have been classified into three major groups - Zn-CDF, Mn-CDF and Zn/Fe-CDF - however, the selectivity of most of the MTPs has not been confirmed yet. Cucumber gene CsMTP9 encoding a putative CDF transporter homologous to members of the Mn-CDF cluster is expressed exclusively in roots. The relative abundance of CsMTP9 transcript and protein in roots is significantly increased under Mn excess and Cd. Immunolocalization with specific antibodies revealed that CsMTP9 is a plasma membrane transporter that localizes to the inner PM domain of root endodermal cells. The plasma membrane localization of CsMTP9 was confirmed by the expression of the fusion proteins of GFP (green fluorescent protein) and CsMTP9 in yeast and protoplasts prepared from Arabidopsis cells. In yeast, CsMTP9 transports Mn2+ and Cd2+ via a proton-antiport mechanism with an apparent Km values of approximately 10 μm and 2.5 μm for Mn2+ and Cd2+, respectively. In addition, CsMTP9 expression in yeast rescues the Mn- and Cd-hypersensitive phenotypes through the enhanced efflux of Mn2+ and Cd2+ from yeast cells. Similarly, the overexpression of CsMTP9 in A. thaliana confers increased resistance of plants to Mn excess and Cd but not to other heavy metals and leads to the enhanced translocation of manganese and cadmium from roots to shoots. These findings indicate that CsMTP9 is a plasma membrane H+-coupled Mn2+ and Cd2+ antiporter involved in the efflux of manganese and cadmium from cucumber root cells by the transport of both metals from endodermis into vascular cylinder. Significance Statement Cation diffusion facilitator (CDF) membrane transporters are important for heavy metal ion transport and in plants such transporters are termed Metal Tolerance Proteins (MTP). Here we characterize a root-specific MTP from cucumber and show that its overexpression confers increased resistance to manganese and cadmium, likely by enhancing transport of these heavy metals from roots to shoots.