[Show abstract][Hide abstract] ABSTRACT: Main conclusion:
The heterologous expression of AtPCS1 in tobacco plants exposed to arsenic plus cadmium enhances phytochelatin levels, root As/Cd accumulation and pollutants detoxification, but does not prevent root cyto-histological damages. High phytochelatin (PC) levels may be involved in accumulation and detoxification of both cadmium (Cd) and arsenic (As) in numerous plants. Although polluted environments are frequently characterized by As and Cd coexistence, how increased PC levels affect the adaptation of the entire plant and the response of its cells/tissues to a combined contamination by As and Cd needs investigation. Consequently, we analyzed tobacco seedlings overexpressing Arabidopsis phytochelatin synthase1 gene (AtPCS1) exposed to As and/or Cd, to evaluate the levels of PCs and As/Cd, the cyto-histological modifications of the roots and the Cd/As leaf extrusion ability. When exposed to As and/or Cd the plants overexpressing AtPCS1 showed higher PC levels, As plus Cd root accumulation, and detoxification ability than the non-overexpressing plants, but a blocked Cd-extrusion from the leaf trichomes. In all genotypes, As, and Cd in particular, damaged lateral root apices, enhancing cell-vacuolization, causing thinning and stretching of endodermis initial cells. Alterations also occurred in the primary structure region of the lateral roots, i.e., cell wall lignification in the external cortex, cell hypertrophy in the inner cortex, crushing of endodermis and stele, and nuclear hypertrophy. Altogether, As and/or Cd caused damage to the lateral roots (and not to the primary one), with such damage not counteracted by AtPCS1 overexpression. The latter, however, positively affected accumulation and detoxification to both pollutants, highlighting that Cd/As accumulation and detoxification due to PCS1 activity do not reduce the cyto-histological damage.
[Show abstract][Hide abstract] ABSTRACT: Phytoplasmas, biotrophic wall-less prokaryotes, only reside in sieve elements of their host plants. The essentials of the intimate interaction between phytoplasmas and their hosts are poorly understood, which calls for research on potential ultrastructural modifications. We investigated modifications of the sieve-element ultrastructure induced in tomato plants by ‘Candidatus Phytoplasma solani’, the pathogen associated with the stolbur disease. Phytoplasma infection induces a drastic re-organization of sieve-element substructures including changes in plasma membrane surface and distortion of the sieve-element reticulum. Observations of healthy and stolbur-diseased plants provided evidence for the emergence of structural links between sieve-element plasma membrane and phytoplasmas. One-sided actin aggregates on the phytoplasma surface also inferred a connection between phytoplasma and sieve-element cytoskeleton. Actin filaments displaced from the sieve-element mictoplasm to the surface of the phytoplasmas in infected sieve elements. Expression analysis revealed a decrease of actin and an increase of ER-resident chaperone luminal binding protein (BiP) in midribs of phytoplasma-infected plants. Collectively, the studies provided novel insights into ultrastructural responses of host sieve elements to phloem-restricted prokaryotes.
Full-text · Article · Aug 2015 · Frontiers in Plant Science
[Show abstract][Hide abstract] ABSTRACT: Over time, anthropogenic activities have led to severe cadmium (Cd) and arsenic (As) pollution in several environments. Plants inhabiting metal(loid)-contaminated areas should be able to sequester and detoxify these toxic elements as soon as they enter roots and leaves. We postulated here that an important role in protecting plants from excessive metal(loid) accumulation and toxicity might be played by arbuscular mycorrhizal (AM) fungi. In fact, human exploitation of plant material derived from Cd- and As-polluted environments may lead to a noxious intake of these toxic elements; in particular, a possible source of Cd and As for humans is given by cigarette and cigar smoke. We investigated here the role of AM fungus Funneliformis mosseae (T.H. Nicolson & Gerd.) C. Walker & A. Schüßler in protecting Nicotiana tabacum L. (cv. Petit Havana) from the above-mentioned meta(loid) stress. Our findings proved that the AM symbiosis is effective in increasing the plant tissue content of the antioxidant glutathione (GSH), in influencing the amount of metal(loid)-induced chelators as phytochelatins, and in reducing the Cd and As content in leaves and roots of adult tobacco plants. These results might also prove useful in improving the quality of commercial tobacco, thus reducing the risks to human health due to inhalation of toxic elements contained in smoking products.
Full-text · Article · Jan 2015 · Plant Physiology and Biochemistry
[Show abstract][Hide abstract] ABSTRACT: Lunularia cruciata occupies a very basal position in the phylogenetic tree of liverworts, which in turn have been recognized as a very early clade of land plants. It would therefore seem appropriate to take L. cruciata as the starting point for investigating character evolution in the metal(loid) response of land plants. One of the strongest evolutionary pressures for land colonization by plants has come from potential access to much greater amounts of nutritive ions from surface rocks. This might have resulted in the need to precisely regulate trace element homeostasis and to minimize the risk of exposure to toxic concentrations of certain metals, prompting the evolution of a number of response mechanisms, such as synthesis of phytochelatins, metal(loid)-binding thiol-peptides. Accordingly, if the ability to synthesize phytochelatins and the occurrence of an active phytochelatin synthase are traits present in a basal liverwort, and have been even reinforced in “modern” tracheophytes, e.g. Arabidopsis thaliana, then such traits would presumably have played an essential role in plant fitness over time. Hence, we demonstrated here that: 1) L. cruciata compartmentalizes cadmium in the vacuoles of the phototosynthetic parenchyma by means of a phytochelatin-mediated detoxification strategy, and possesses a phytochelatin synthase that is activated by cadmium and homeostatic concentrations of iron(II) and zinc; 2) A. thaliana phytochelatin synthase displays a higher and broader response to several metal(loid)s [namely: cadmium, iron(II), zinc, copper, mercury, lead, arsenic(III)] than the phytochelatin synthase of L. cruciata.
[Show abstract][Hide abstract] ABSTRACT: The grapevine (Vitis vinifera) disease caused by Bois Noir (BN) phytoplasma, a plant phloem-limited pathogens belonging to the class Mollicutes, is an important concern to European vinery economy. Phytoplasmas have a broad range of plant hosts among monocots and dicots and diseases of many important crops have already been associated with these pathogens. Leaf curling and discoloration, uneven or total lack of lignification of canes, flower abortion and berry withering are the typical symptoms of phytoplasma infection in grapevine. An interesting aspect of the pathogen–plant interaction is “recovery” phenomenon, a spontaneous remission of symptoms in previously symptomatic plants. In this study, we are using 2-DE and Western Blot analysis to explore the proteome changes in leaves and midribs of V. vinifera cv. 'Chardonnay' naturally infected by BN, with particular attention to recovered plants.
[Show abstract][Hide abstract] ABSTRACT: A recent study by our group demonstrated that bryophytes - a paraphyletic group which includes liverworts (Marchantiophyta), mosses (Bryophyta) and hornworts (Anthocerotophyta) - have the capability to produce phytochelatins (PCs) in response to Cd, and that they possess a constitutive and functional phytochelatin synthase (PCS). Amongst bryophytes, liverworts occupy an ideal basal phylogenetic position for reconstructing ancient evolutionary changes that occurred over time. Notably, the early Devonian Rhynie Chert of NE Scotland (ca. 396 ± 12 Mya) contained the earliest bryophytic flora and featured high levels of Hg, Zn, Au, the metalloids As and Sb, etc., all PC-inducers to varying extents, at least in higher plants. Accordingly, if the PC production capability and a functional PCS were present even in the earliest land plants (such as ancient liverworts) and have been maintained until today, it might be postulated that PCs and PCS would have played a positive role for plant fitness in metal(loid)-containing palaeoenvironments over the course of time. Thus, in order to identify events of an ancestral nature in the basalmost land plants, we hypothesized here that gametophytes of Lunularia cruciata (Lunulariales, Lunulariaceae), an early divergent clade of thalloid liverworts, would: 1) possess a metal(loid)-responsive PCS activated by Cd and other elements, such as Hg, Pb, Zn, Cu, As, and Sb, which are documented to be present both in the abovementioned palaeoenvironments, and in the present-day environments; 2) compartmentalize the excess of certain metals, i.e. Cd and Zn, in the vacuoles, by means of a PC-mediated detoxification strategy.
[Show abstract][Hide abstract] ABSTRACT: Bryophytes, a paraphyletic group which includes liverworts, mosses, and hornworts, have been stated as land plants that under metal stress (particularly cadmium) do not synthesize metal-binding peptides such as phytochelatins. Moreover, very little information is available to date regarding phytochelatin synthesis in charophytes, postulated to be the direct ancestors of land plants, or in lycophytes, namely very basal tracheophytes. In this study, it was hypothesized that basal land plants and charophytes have the capability to produce phytochelatins and possess constitutive and functional phytochelatin synthases. To verify this hypothesis, twelve bryophyte species (six liverworts, four mosses, and two hornworts), three charophytes, and two lycophyte species were exposed to 0-36 μM cadmium for 72h, and then assayed for: (i) glutathione and phytochelatin quali-quantitative content by HPLC and mass spectrometry; (ii) the presence of putative phytochelatin synthases by western blotting; and (iii) in vitro activity of phytochelatin synthases. Of all the species tested, ten produced phytochelatins in vivo, while the other seven did not. The presence of a constitutively expressed and functional phytochelatin synthase was demonstrated in all the bryophyte lineages and in the lycophyte Selaginella denticulata, but not in the charophytes. Hence, current knowledge according to phytochelatins have been stated as being absent in bryophytes was therefore confuted by this work. It is argued that the capability to synthesize phytochelatins, as well as the presence of active phytochelatin synthases, are ancestral (plesiomorphic) characters for basal land plants.
Full-text · Article · Jan 2014 · Journal of Experimental Botany
[Show abstract][Hide abstract] ABSTRACT: Bryophytes, namely liverworts, mosses and hornworts, have been stated as basal land
plants that, under metal stress (particularly cadmium), do not synthesize metal-binding
peptides such as phytochelatins. We hypothesized instead that a “phytochelatin production
capability” may be present also in these plants. For verifying such hypothesis, twelve
bryophyte species (six liverworts, Conocephalum conicum, Marchantia polymorpha,
Pellia epiphylla, Radula complanata, Aneura pinguis and Scapania undulata; four mosses,
Sphagnum palustre, Polytrichastrum formosum, Hypnum cupressiforme and Fontinalis
antipyretica; and two hornworts, Anthoceros punctatus and Phaeoceros laevis) were
grown with or without 36 microM cadmium for 72 h, and then assayed for phytochelatin
quali-quantitative content by post-column-derivatized HPLC followed by mass and mass/
mass spectrometry. In addition, western blot analyses for immunodetection of putative
phytochelatin synthases were performed in C. conicum, S. palustre, P. laevis and, as a
negative control, in Physcomitrella patens, whose genome is completely sequenced. Of
the twelve species tested, six produced phytochelatins (four liverworts, one moss, one
hornwort), whereas six did not. Not least, the presence of an endogenous, constitutivelyexpressed,
phytochelatin synthase was also found in some species. Hence, bryophytes do
produce phytochelatins, thus confuting the current knowledge according to which these
thiol-peptides are absent in such plants; besides, phytochelatins were mainly detected
in stem lineages, whereas they appeared to be less abundant (or absent) in crown taxa.
This suggests that phytochelatin synthesis, as well as the presence of a constitutive
phytochelatin synthase, is an ancestral character for basal land plants.
[Show abstract][Hide abstract] ABSTRACT: Very few studies have provided information about the effects of cadmium (Cd) at histoanatomical and ultrastructural levels, along with potential localization of the metal in planta. In particular, from this standpoint, almost nothing is known in Daucus carota L. (carrot), a particularly important species for in vitro and in vivo functional investigations. In this work we hypothesized that 36 μM Cd, supplied for 1, 2, 3, 4, 7 and 14 days to 30-day-old in vitro-cultured plants, might induce an early acclimation, but a final collapse of roots and leaves. In fact, as a general feature, a biphasic root response to Cd stress actually took place: in the first phase (1-4 days of Cd exposure), the cytological and functional events observed - by light microscopy, TEM, epifluorescence, as well as by the time-course of thiol-peptide compounds - can be interpreted as acclimatory responses aimed at diminishing the movement of Cd across the root. The second phase (from 4 to 14 days of Cd exposure) was instead characterized by cell hypertrophy, cell-to-cell separation events, increase in α-β-γ-tocopherol levels and, not least, endocytogenic processes, coupled with a dramatic drop in the amount of thiol-peptide compounds. These events led to a progressive root collapse, even if they did not ingenerate macro/microscopic injury symptoms in leaf blades and petioles.
[Show abstract][Hide abstract] ABSTRACT: Indian mustard (Brassica juncea L. Czern.) tolerates high concentrations of heavy metals and is a promising species for the purpose of phytoextraction of cadmium (Cd) from metal-contaminated soils. This work investigates the extent to which antioxidant and metal sequestering mechanisms are responsible for this tolerance. To this end, seedlings of Indian mustard were grown for 7 days in 0, 50 or 200 μM Cd. Increasing Cd concentrations led to a progressive Cd accumulation in roots and shoots, accompanied by an organ-dependent alteration in mineral uptake, and a decrease in root/shoot length and fresh/dry weight. Cd negatively affected chlorophyll and carotenoid contents and activated the xanthophyll cycle, suggesting the need to protect the photosynthetic apparatus from photoinhibition. Shoots seemed to be less efficient than roots in ROS scavenging, as indicated by the different response to Cd stress shown by peroxidase and catalase activities and, solely with regard to the highest Cd concentration, by ascorbate level. Such a different antioxidant capacity might at least partly explain differences in the trend of lipid peroxidation observed in the two organs. Moreover, in both roots and shoots, glutathione and phytochelatin content markedly increased under Cd stress, regardless of the metal concentration involved.
No preview · Article · May 2012 · Plant Physiology and Biochemistry
[Show abstract][Hide abstract] ABSTRACT: Both Fe deficiency and Cd exposure induce rapid changes in the S nutritional requirement of plants. The aim of this work was to characterize the strategies adopted by plants to cope with both Fe deficiency (release of phytosiderophores) and Cd contamination [production of glutathione (GSH) and phytochelatins] when grown under conditions of limited S supply. Experiments were performed in hydroponics, using barley plants grown under S sufficiency (1.2 mM sulphate) and S deficiency (0 mM sulphate), with or without Fe(III)-EDTA at 0.08 mM for 11 d and subsequently exposed to 0.05 mM Cd for 24 h or 72 h. In S-sufficient plants, Fe deficiency enhanced both root and shoot Cd concentrations and increased GSH and phytochelatin levels. In S-deficient plants, Fe starvation caused a slight increase in Cd concentration, but this change was accompanied neither by an increase in GSH nor by an accumulation of phytochelatins. Release of phytosiderophores, only detectable in Fe-deficient plants, was strongly decreased by S deficiency and further reduced after Cd treatment. In roots Cd exposure increased the expression of the high affinity sulphate transporter gene (HvST1) regardless of the S supply, and the expression of the Fe deficiency-responsive genes, HvYS1 and HvIDS2, irrespective of Fe supply. In conclusion, adequate S availability is necessary to cope with Fe deficiency and Cd toxicity in barley plants. Moreover, it appears that in Fe-deficient plants grown in the presence of Cd with limited S supply, sulphur may be preferentially employed in the pathway for biosynthesis of phytosiderophores, rather than for phytochelatin production.
Full-text · Article · Nov 2011 · Journal of Experimental Botany
[Show abstract][Hide abstract] ABSTRACT: The hypothesis that a daily water supply allows a lichen to endure the negative effects of environmental concentrations of NO(x) and O(3) was tested with a transplant experiment. Five groups (0, A-D) of Flavoparmelia caperata samples derived from the same thalli were used for destructive, pre-exposure measurements (0), or exposed for 5 weeks in the rural collection site (A), and in a urban site with high levels of NO(x) and O(3) (B-D). Two groups (C, D) were daily watered half an hour before the daily peak of NO(x) (C), and O(3) (D). The comparison between pre- and post-exposure measurements of stress biomarkers revealed that the different thallus hydration regime modified the pollution tolerance as well as the physiology of the exposed samples. The non-watered group B suffered an evident decrease in F(v)/F(m) and reduced glutathione, but increased ion leakage, whereas the watered groups C and D showed only decreased non-photosynthetic-quenching, possibly derived from NO(x) exposure. Ozone, which was higher in the rural than in the urban site, did not significantly affect the lichen metabolism. Our results re-open the discussion on the so-called "drought hypothesis", which suggests that the lichen desert observed in urban areas of central and eastern Europe is more a matter of dry microclimate than of air pollution.
[Show abstract][Hide abstract] ABSTRACT: Previous studies demonstrated that expression of the Arabidopsis phytochelatin (PC) biosynthetic gene AtPCS1 in Nicotiana tabacum plants increases the Cd tolerance in the presence of exogenous glutathione (GSH). In this paper, the Cd tolerance of Arabidopsis plants over-expressing AtPCS1 (AtPCSox lines) has been analysed and the differences between Arabidopsis and tobacco are shown. Based on the analysis of seedling fresh weight, primary root length, and alterations in root anatomy,
evidence is provided that, at relatively low Cd concentrations, the Cd tolerance of AtPCSox lines is lower than the wild type,
while AtPCS1 over-expressing tobacco is more tolerant to Cd than the wild type. At higher Cd concentrations, Arabidopsis AtPCSox seedlings are more tolerant to Cd than the wild type, while tobacco AtPCS1 seedlings are as sensitive as the wild type. Exogenous GSH, in contrast to what was observed in tobacco, did not increase
the Cd tolerance of AtPCSox lines. The PC content in wild-type Arabidopsis at low Cd concentrations is more than three times higher than in tobacco and substantial differences were also found in the
PC chain lengths. These data indicate that the differences in Cd tolerance and in its dependence on exogenous GSH between
Arabidopsis and tobacco are due to species-specific differences in the endogenous content of PCs and GSH and may be in the relative abundance
of PCs of different length.
Full-text · Article · Aug 2011 · Journal of Experimental Botany