PuHSFA4a Enhances Tolerance To Excess Zinc by Regulating Reactive Oxygen Species Production and Root Development in <i>Populus</i>

Abstract

Zinc (Zn) is an essential micronutrient but in excess is highly toxic to plants. Plants regulate Zn homeostasis and withstand excess Zn through various pathways; these pathways are generally tightly regulated by a specific set of genes. However, the transcription factors involved in excess Zn tolerance have yet to be identified. Here, we characterized a <i>Populus ussuriensis</i> heat shock transcription factor A4a (PuHSFA4a) that acts as a positive regulator of excess Zn tolerance in <i>P</i> <i>ussuriensis</i> We used overexpression (<i>PuHSFA4a</i>-OE) and chimeric dominant repressor (<i>PuHSFA4a</i>-<i>SRDX</i>) lines to identify the targets of <i>PuHSFA4a</i> <i>PuHSFA4a</i> transcription is specifically induced in roots by high Zn. Overexpression of <i>PuHSFA4a</i> conferred excess Zn tolerance and a dominant repressor version of <i>PuHSFA4a</i> increased excess Zn sensitivity in <i>P</i> <i>ussuriensis</i> by regulating the antioxidant system in roots. PuHSFA4a coordinately activates genes related to abiotic stress responses and root development and directly binds to the promoter regions of glutathione-s-transferase U17 (<i>PuGSTU17</i>) and phospholipase A₂ (<i>PuPLA₂</i> ). <i>PuGSTU17</i> overexpression significantly increased GST activity and reduced reactive oxygen species levels in roots while <i>PuGSTU17</i>-RNA interference lines exhibited the opposite phenotype. Furthermore, <i>PuPLA₂</i> overexpression promoted root growth under high Zn stress. Taken together, we provide evidence that <i>PuHSFA4a</i> coordinately activates the antioxidant system and root development-related genes and directly targets <i>PuGSTU17</i> and <i>PuPLA</i>, thereby promoting excess Zn tolerance in <i>P</i> <i>ussuriensis</i> roots.