Proteasome targeting of proteins in Arabidopsis leaf mesophyll, epidermal and vascular tissues.

Julia Svozil,Wilhelm Gruissem,Katja Baerenfaller

doi:10.3389/fpls.2015.00376

Julia Svozil, Wilhelm Gruissem + Show 1 more

Open Access

https://doi.org/10.3389/fpls.2015.00376

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Abstract

Protein and transcript levels are partly decoupled as a function of translation efficiency and protein degradation. Selective protein degradation via the Ubiquitin-26S proteasome system (UPS) ensures protein homeostasis and facilitates adjustment of protein abundance during changing environmental conditions. Since individual leaf tissues have specialized functions, their protein composition is different and hence also protein level regulation is expected to differ. To understand UPS function in a tissue-specific context we developed a method termed Meselect to effectively and rapidly separate Arabidopsis thaliana leaf epidermal, vascular and mesophyll tissues. Epidermal and vascular tissue cells are separated mechanically, while mesophyll cells are obtained after rapid protoplasting. The high yield of proteins was sufficient for tissue-specific proteome analyses after inhibition of the proteasome with the specific inhibitor Syringolin A (SylA) and affinity enrichment of ubiquitylated proteins. SylA treatment of leaves resulted in the accumulation of 225 proteins and identification of 519 ubiquitylated proteins. Proteins that were exclusively identified in the three different tissue types are consistent with specific cellular functions. Mesophyll cell proteins were enriched for plastid membrane translocation complexes as targets of the UPS. Epidermis enzymes of the TCA cycle and cell wall biosynthesis specifically accumulated after proteasome inhibition, and in the vascular tissue several enzymes involved in glucosinolate biosynthesis were found to be ubiquitylated. Our results demonstrate that protein level changes and UPS protein targets are characteristic of the individual leaf tissues and that the proteasome is relevant for tissue-specific functions.

Highlights

Plant organs are composed of different tissues that are specialized for particular biological processes and the functionality of the organ is the sum of the functions of each of its tissue types
In the tissue-type specific protein extracts of leaves from the KC464 line, which expresses green fluorescent protein (GFP) exclusively in the epidermis, the GFP protein was exclusively found in the epidermal protein extract but not in the protein extract of vasculature and mesophyll tissues (Figure 1A)
In the JR11-2 line backcrossed with Col-0, GFP is expressed in the spongy mesophyll and GFP was detected only the mesophyll protein extract, confirming that both vasculature and epidermal protein extracts do not contain mesophyll proteins (Figure 1E)

Summary

Introduction

Plant organs are composed of different tissues that are specialized for particular biological processes and the functionality of the organ is the sum of the functions of each of its tissue types. Just as each Arabidopsis organ has its own functional proteome map (Baerenfaller et al, 2008) we expect that individual tissues have specific protein compositions for their specific functions. The main leaf tissue types are epidermis, mesophyll and vasculature. In Arabidopsis, the epidermis is composed of stomata and trichomes that are embedded in the single cell layer of pavement cells, which are covered with the waxy cuticle. The adaxial and abaxial epidermal tissues enclose the palisade and spongy mesophyll, Proteasome targets in different leaf tissues which represent the main photosynthetic capacity of the leaf. Embedded in the mesophyll tissue is the vascular tissue, which consists of phloem, xylem and cambial cells (Tsukaya, 2002)

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