A proteomics approach to investigate the process of <scp>Z</scp>n hyperaccumulation in <i><scp>N</scp>occaea caerulescens</i> (<scp>J</scp> & <scp>C</scp>. <scp>P</scp>resl) <scp>F</scp>.<scp>K</scp>. <scp>M</scp>eyer

<jats:title>Summary</jats:title><jats:p>Zinc (<jats:styled-content style="fixed-case">Z</jats:styled-content>n) is an essential trace element in all living organisms, but is toxic in excess. Several plant species are able to accumulate <jats:styled-content style="fixed-case">Z</jats:styled-content>n at extraordinarily high concentrations in the leaf epidermis without showing any toxicity symptoms. However, the molecular mechanisms of this phenomenon are still poorly understood. A state‐of‐the‐art quantitative 2<jats:styled-content style="fixed-case">D</jats:styled-content> liquid chromatography/tandem mass spectrometry (2<jats:styled-content style="fixed-case">D</jats:styled-content>‐<jats:styled-content style="fixed-case">LC</jats:styled-content>‐<jats:styled-content style="fixed-case">MS</jats:styled-content>/<jats:styled-content style="fixed-case">MS</jats:styled-content>) proteomics approach was used to investigate the abundance of proteins involved in <jats:styled-content style="fixed-case">Z</jats:styled-content>n hyperaccumulation in leaf epidermal and mesophyll tissues of <jats:italic><jats:styled-content style="fixed-case">N</jats:styled-content>occaea caerulescens</jats:italic>. Furthermore, the <jats:styled-content style="fixed-case">Z</jats:styled-content>n speciation <jats:italic>in planta</jats:italic> was analyzed by a size‐exclusion chromatography/inductively coupled plasma mass spectrometer (<jats:styled-content style="fixed-case">SEC</jats:styled-content>‐<jats:styled-content style="fixed-case">ICP</jats:styled-content>‐<jats:styled-content style="fixed-case">MS</jats:styled-content>) method, in order to identify the <jats:styled-content style="fixed-case">Z</jats:styled-content>n‐binding ligands and mechanisms responsible for <jats:styled-content style="fixed-case">Z</jats:styled-content>n hyperaccumulation. Epidermal cells have an increased capability to cope with the oxidative stress that results from excess <jats:styled-content style="fixed-case">Z</jats:styled-content>n, as indicated by a higher abundance of glutathione <jats:italic><jats:styled-content style="fixed-case">S</jats:styled-content></jats:italic>‐transferase proteins. A <jats:styled-content style="fixed-case">Z</jats:styled-content>n importer of the <jats:styled-content style="fixed-case">ZIP</jats:styled-content> family was more abundant in the epidermal tissue than in the mesophyll tissue, but the vacuolar <jats:styled-content style="fixed-case">Z</jats:styled-content>n transporter <jats:styled-content style="fixed-case">MTP</jats:styled-content>1 was equally distributed. Almost all of the <jats:styled-content style="fixed-case">Z</jats:styled-content>n located in the mesophyll was stored as <jats:styled-content style="fixed-case">Z</jats:styled-content>n–nicotianamine complexes. In contrast, a much lower proportion of the <jats:styled-content style="fixed-case">Z</jats:styled-content>n was found as <jats:styled-content style="fixed-case">Z</jats:styled-content>n–nicotianamine complexes in the epidermis. However, these cells have higher concentrations of malate and citrate, and these organic acids are probably responsible for complexation of most epidermal <jats:styled-content style="fixed-case">Z</jats:styled-content>n. Here we provide evidence for a cell type‐specific adaptation to excess <jats:styled-content style="fixed-case">Z</jats:styled-content>n conditions and an increased ability to transport <jats:styled-content style="fixed-case">Z</jats:styled-content>n into the epidermal vacuoles.</jats:p>