XB-ART-56235Front Plant Sci January 1, 2019; 10 988.
Silicon Uptake and Localisation in Date Palm (Phoenix dactylifera) - A Unique Association With Sclerenchyma.
Date palm (Phoenix dactylifera) can accumulate as much as 1% silicon (Si), but not much is known about the mechanisms inherent to this process. Here, we investigated in detail the uptake, accumulation and distribution of Si in date palms, and the phylogeny of Si transporter genes in plants. We characterized the PdNIP2 transporter following heterologous expression in Xenopus oocytes and used qPCR to determine the relative expression of Si transporter genes. Silicon accumulation and distribution was investigated by light microscopy, scanning electron microscopy coupled with X-ray microanalysis and Raman microspectroscopy. We proved that PdNIP2-1 codes for a functional Si-permeable protein and demonstrated that PdNIP2 transporter genes were constitutively expressed in date palm. Silicon aggregates/phytoliths were found in specific stegmata cells present in roots, stems and leaves and their surfaces were composed of pure silica. Stegmata were organized on the outer surface of the sclerenchyma bundles or associated with the sclerenchyma of the vascular bundles. Phylogenetic analysis clustered NIP2 transporters of the Arecaceae in a sister position to those of the Poaceae. It is suggested, that Si uptake in date palm is mediated by a constitutively expressed Si influx transporter and accumulated as Si aggregates in stegmata cells abundant in the outer surface of the sclerenchyma bundles (fibers).
PubMed ID: 31456812
PMC ID: PMC6701203
Article link: Front Plant Sci
Article Images: [+] show captions
|FIGURE 1. Silicon concentration in vegetative organs of Phoenix dactylifera cultivated in hydroponics, perlite or soil. The plants cultivated in perlite were 1-year-old in comparison to the well-developed, 10-year-old plants grown in a soil. Different letters indicate significant differences between the treatments at 0.05 level. Values are means (n = 4) ± standard deviation.|
|FIGURE 2. The anatomy of individual root types in P. dactylifera with fiber bands and stegmata – specialized Si-accumulating cells. (A) Seminal root. (B,C) Lateral roots. (D–J) Adventitious roots in transverse (D–H) and longitudinal sections (I,J). Paraffin embedded sections stained with alcian blue/safranin (A,B,D,E,H–J) or with Basic fuchsin (F), unstained – autofluorescence (C), and hand sections stained with Fluorol yellow in UV light (G). (A) Seminal root of 2-month-old seedling covered by rhizodermis (epidermis) (e) and composed hypodermis. Mid-cortex typically develops an extensive aerenchyma (aer). Polyarch vascular cylinder (vc) with 11 alternating xylem and phloem poles, several late metaxylem vessels are shifted centripetally and sclerified pith is located in the center. No fiber bands are present at this stage of development. (B,C) Lateral roots structurally resemble the seminal root, with exception of fiber bands (white arrowheads) regularly scattered in the cortex. Stegmata cells, adjacent to the fiber bands, are not visible at this magnification. Rhizodermis (e) and composed hypodermis (hyp) are formed by several layers of cells with varying cell wall thicknesses. Extensive aerenchyma (aer) occupies the mid-cortex. Endodermis (en) with thick U-shaped inner tangential walls. Broad late metaxylem vessels (lx) are shifted centripetally from the xylem poles formed by early metaxylem vessels (ex). (D–J) Adventitious roots of adult plants are characterised by a multitude of fiber bands (white arrowheads) scattered in the mid-cortex. Proportionally to the age/thickness of the adventitious roots, the number of fiber bands counts from dozens (F,G) to hundreds (D,E,H). Composed hypodermis (hyp) is formed by several layers of exodermis with suberized cell walls (G,H). Outer cortex is located internally to the hypodermis, composed of several layers of sclerenchyma. Many cells in the peripheral tissues contain tannins (tc). Polyarch vascular cylinder with several dozens of alternating xylem (ex) and phloem (ph) poles is surrounded by thick-walled endodermis (en). An additional circle of late metaxylem (lx) vessels is present centripetally from xylem poles. In thick roots, the pith (p) might form a central cavity. (E) Stegmata with silica phytoliths (red arrowheads) are attached to the surface of fiber bands and are clearly visible in longitudinal sections of roots (I,J).|
|FIGURE 3. Structure of stem apex (A), young leaf (B), leaflet blade (C–G), leaf petiole (H,I), and leaf sheath (J–L). Stegmata in leaves are associated with both fiber bands and vascular sclerenchyma. (A) Stereomicroscope image of the shoot apex showing the shoot apical meristem (sam), leaf primordia (lp), vascular bundles (vb), and fiber bands (white arrowheads). (B) Unstained paradermal section of a young leaf from a date palm seedling showing epidermal cells, stoma (st), and subepidermally occurring stegmata (red arrowheads) attached to the surface of a fiber band. (C–G) Adult leaflet from a 10-year old. Unstained sections (C,E) show epidermal and hypodermal layers at the leaf surface, mesophyll with vascular bundles (vb), and fiber bands (white arrowheads) occurring both adaxially and abaxially. In the central part of the leaflet the mid vein is absent (E) and expanding tissue of large parenchyma cells is present adaxially (exp). In the opposite-abaxial part, two large fiber bands are developed. Stegmata adjacent to the fiber bands and sclerenchyma sheaths of vascular bundles can be seen in high magnification (D) and more clearly in longitudinal sections (F,G). Alcian blue/safranin stained samples (D,F,G). (H,I) Leaf petiole and (J–L) leaf sheaths of adult 10-year-old plant in unstained cross-sections are shown either under white light (H,J,L) or UV irradiation (I). (J,K) Phloroglucinol-HCl staining visualizing cell wall lignification. Leaf petiole and leaf sheath are covered by a single layer epidermis and lignified hypodermis (I,J). Mesophyll is composed of chlorenchyma (H,I) and parenchymatous ground tissue. Peripherally present fiber bands (white arrowheads) and large sclerenchyma sheaths of vascular bundles are accompanied by stegmata, as seen in high magnification (F,G,K,L). The ground tissue cells usually contain a number of starch grains (sg).|
|FIGURE 4. Scanning electron microscopy images of various P. dactylifera tissues with corresponding maps showing the distribution of Si (violet color). (A,B) Cross section of an adventitious root showing detail of a fiber band (white arrowhead) with adjacent stegmata cells containing Si phytoliths (red arrowheads). Multiple phytoliths are not visible in (A), though detected by EDX (B). (C,D) Longitudinal section through the fiber band (white arrowhead) in an adventitious root. Cell walls of several stegmata cells are disrupted, uncovering Si phytoliths (red arrowheads). (E,F) Cross section of a leaf showing the presence of Si phytoliths in stegamata cells associated with vascular bundles (vb) and fiber bands (white arrowheads). A detail on a stegma (red arrowhead) associated with the vascular bundle sclerenchyma (scl). (G,H) A surface view on a fiber band (white arrowhead) with a dense net of adjacent stegmata. Cell walls of multiple stegmata are disrupted, uncovering Si phytoliths (red arrowheads).|
|FIGURE 5. EDX and Raman analyses of the Si phytoliths. (A) Scanning electron micrograph showing three adjacent stegmata with disrupted cell walls exposing the Si phytoliths. (B) Detail of an exposed phytolith used in EDX analysis. (C) Dark-field microscopy image of isolated Si phytoliths used in Raman analysis. (D) A representative spectrum from EDX analysis of a Si phytolith demonstrating the dominance of Si and O as its main chemical constituents. (E) Comparison of Raman spectra collected from isolated silica phytoliths [shown in panel (C)], hydrated silica gel, and opal.|
|FIGURE 6. Alignment of amino acid sequences of silicon influx transporters (NIP2s) in various plant species with highlighted NPA motifs (red color) and the G-S-G-R Ar/R selectivity filter (green color) (A). Prediction of the 3D structure of PdNIP2-1 (B) and PdNIP2-2 (C) proteins. Prediction of transmembrane domains of PdNIP2-1 (D) and PdNIP2-2 (E) proteins.|
|FIGURE 7. Phylogeny of silicon influx transporters (NIP2s) of monocotyledonous and dicotyledonous plants. Posterior probabilities for Bayesian inference and bootstrap values for maximum likelihood were mapped onto the 50%-majority rule consensus tree. The scale bar indicates two substitutions per ten hundred amino acid positions.|
|FIGURE 8. (A) Silicon influx transport activity of PdNIP2-1 from date palm evaluated at two different time points in Xenopus oocyte assays. Oocytes injected with OsLsi1 from rice, or water were used as positive and negative controls, respectively. Values are means ± standard deviation. Different letters indicate significant differences in the same time point. The relative transcript level of PdNIP2-1 (B) and PdNIP2-2 (C) genes in roots of hydroponically grown date palm seedlings in the Si– treatment (orange line) and the Si+ treatment (blue line) from the third to the seventh day of cultivation. Gene expression for the control was set as 1.0. Statistically significant differences between control and treated plants were analyzed by Student’s t test and are denoted as ∗P < 0.05. Values are means ± standard deviation. The mean values are based on three technical and three biological replicates.|
References [+] :
Bokor, Expression of Genes for Si Uptake, Accumulation, and Correlation of Si with Other Elements in Ionome of Maize Kernel. 2019, Pubmed