Rice phytochelatin synthases Os<scp>PCS</scp>1 and Os<scp>PCS</scp>2 make different contributions to cadmium and arsenic tolerance

<jats:title>Abstract</jats:title><jats:p>Cadmium (Cd) and arsenic (As) pollution in paddy soil and their accumulation in rice (<jats:italic>Oryza sativa</jats:italic>) pose serious threats to human health. Rice internally detoxifies these toxic metal and metalloid to some extent, resulting in their accumulation within the edible parts. However, the mechanisms of Cd and As detoxification in rice have been poorly elucidated. Plants synthesize thiol‐rich metal‐chelating peptides, termed phytochelatins (<jats:styled-content style="fixed-case">PC</jats:styled-content>s). We characterized rice <jats:styled-content style="fixed-case">PC</jats:styled-content> synthase (<jats:styled-content style="fixed-case">PCS</jats:styled-content>) and investigated its contribution to Cd and As tolerance in rice. We identified two <jats:italic><jats:styled-content style="fixed-case">PCS</jats:styled-content></jats:italic> homolog genes, <jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>1</jats:italic> and <jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>2</jats:italic>, in the rice genome. The expression of <jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>1</jats:italic> was upregulated by As(<jats:styled-content style="fixed-case">III</jats:styled-content>) stress in the roots but that of <jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>2</jats:italic> was not significantly affected. The expression level of <jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>2</jats:italic> was higher than that of <jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>1</jats:italic> in the shoots and roots. Recombinant Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>1 and Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>2 proteins differed in their metal activation. Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>1 was more strongly activated by As(<jats:styled-content style="fixed-case">III</jats:styled-content>) than by Cd; however, Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>2 was more strongly activated by Cd than by As(<jats:styled-content style="fixed-case">III</jats:styled-content>). Genetically engineered plants having their <jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>2</jats:italic> expression silenced via <jats:styled-content style="fixed-case">RNA</jats:styled-content> interference (<jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>2 </jats:italic><jats:styled-content style="fixed-case">RNA</jats:styled-content>i) contained less <jats:styled-content style="fixed-case">PC</jats:styled-content>s and more glutathione (<jats:styled-content style="fixed-case">GSH</jats:styled-content>), a substrate of <jats:styled-content style="fixed-case">PC</jats:styled-content> synthesis, than wild‐type plants, although there was no significant difference in <jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>1 </jats:italic><jats:styled-content style="fixed-case">RNA</jats:styled-content>i plants. <jats:italic>Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>2 </jats:italic><jats:styled-content style="fixed-case">RNA</jats:styled-content>i plants were sensitive to As(<jats:styled-content style="fixed-case">III</jats:styled-content>) stress, but Cd tolerance was little affected. On the other hand, treatment with buthionine sulfoximine, an inhibitor of <jats:styled-content style="fixed-case">GSH</jats:styled-content> biosynthesis, significantly decreased Cd and As tolerance of rice seedlings. These findings indicate that Os<jats:styled-content style="fixed-case">PCS</jats:styled-content>2 is a major isozyme controlling <jats:styled-content style="fixed-case">PC</jats:styled-content> synthesis, and that <jats:styled-content style="fixed-case">PC</jats:styled-content>s are important for As tolerance in rice. However, <jats:styled-content style="fixed-case">PC</jats:styled-content> synthesis may make a smaller contribution to Cd tolerance in rice, and <jats:styled-content style="fixed-case">GSH</jats:styled-content> plays crucial roles, not only as a substrate of <jats:styled-content style="fixed-case">PC</jats:styled-content> synthesis.</jats:p>