Eukaryotic and Prokaryotic Phytochelatin Synthases Differ Less in Functional Terms Than Previously Thought: A Comparative Analysis of Marchantia polymorpha and Geitlerinema sp. PCC 7407.

Erika Bellini,Lorenza Rugnini,Laura Bruno,Marco Borsò,Luigi Sanità di Toppi,Claudio Varotto

doi:10.3390/plants9070914

Erika Bellini, Lorenza Rugnini + Show 4 more

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https://doi.org/10.3390/plants9070914

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Abstract

This paper reports functional studies on the enzyme phytochelatin synthase in the liverwort Marchantia polymorpha and the cyanobacterium Geitlerinema sp. strain PCC 7407. In vitro activity assays in control samples (cadmium-untreated) showed that phytochelatin synthase was constitutively expressed in both organisms. In the presence of 100 µM cadmium, in both the liverwort and the cyanobacterium, the enzyme was promptly activated in vitro, and produced phytochelatins up to the oligomer PC4. Likewise, in vivo exposure to 10–36 µM cadmium for 6-120 h induced in both organisms phytochelatin synthesis up to PC4. Furthermore, the glutathione (GSH) levels in M. polymorpha were constitutively low (compared with the average content in higher plants), but increased considerably under cadmium stress. Conversely, the GSH levels in Geitlerinema sp. PCC 7407 were constitutively high, but were halved under metal treatments. At odds with former papers, our results demonstrate that, as in M. polymorpha and other plants, the cyanobacterial phytochelatin synthase exposed to cadmium possesses manifest transpeptidasic activity, being able to synthesize phytochelatins with a degree of oligomerization higher than PC2. Therefore, prokaryotic and eukaryotic phytochelatin synthases differ less in functional terms than previously thought.

Highlights

Phytochelatin synthase (PCS) is a γ-glutamylcysteine-dipeptidyl-(trans)peptidase (EC 2.3.2.15)belonging to the “Clan CA” of papain-like cysteine proteases [1,2]
In this paper, we have focused our attention on two photoautotrophic model-organisms, namely the cyanobacterium Geitlerinema sp. strain PCC 7407 and the liverwort Marchantia polymorpha L
Table S1). subsp. ruderalis) and the is constitutively expressed in both theranging liverwort (gametophytes of M. polymorpha cyanobacterium; (2) in the absence of Cd, the PCSs of both organisms are able to produce a basal level of PCn; (3) in the presence of Cd at all concentrations, the PCSs from the two organisms are promptly activated and produce PCn, namely PC2, PC3, and PC4; (4) in both organisms, the increasing Cd concentrations and exposure times result in an increase in PCn levels, PC2, PC3, and PC4 in the liverwort and PC3 in the cyanobacterium; (5) the constitutive levels of GSH in the liverwort, relative to those normally found in higher plants, are quite low, but increase significantly under Cd treatments

Summary

Introduction

Phytochelatin synthase (PCS) is a γ-glutamylcysteine-dipeptidyl-(trans)peptidase (EC 2.3.2.15)belonging to the “Clan CA” of papain-like cysteine proteases [1,2]. Eukaryotic PCSs, plant PCSs, display manifest transpeptidasic activity by catalyzing the cytosolic synthesis of phytochelatin (PCn) thiol-oligopeptides, whose general structure is (γ-glutamate–cysteine)n -glycine, with n ranging from 2 to 5; for this reason, PCn oligomers are named PC2 , PC3 , PC4 , and PC5. The cysteine thiol-groups enable PCn to chelate several metal(loid)s. Plants 2020, 9, 914 and segregate them in the vacuo-lysosomal compartments [2,11], drastically reducing the metal(loid) toxicity. The transpeptidasic activity of PCS is manifested when the enzyme’s catalytic site tightly binds complexes between glutathione in reduced form (GSH) or its direct thiol-derivatives, and metal(loid)s such as cadmium (Cd), lead (Pb), mercury (Hg), arsenic (As), copper (Cu), zinc (Zn), and iron (Fe) [3,11]. The very limited number of studies conducted on cyanobacterial PCS—in particular on Nostoc sp

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