While liming soils is technically feasible, the high levels of lime needed to change soil pH, the limitations of lime penetrating beneath surface soils, and the high cost of transporting the soil additive to countries which lack sources of lime make this practice largely unsustainable, unaffordable and inefficient.The toxic effect of Al on plant development has been reported on for nearly a century and the predominant and immediate consequence of Al toxicity is the inhibition of primary root growth . At the cellular and biochemical level, Al has been found to interrupt cell division and elongation, nutrient uptake, Ca2+ homeostasis, IP3 and hormone signaling, cytoskeletal structure, vesicle trafficking, plasma membrane integrity, and chromatin structure . Due to the ubiquitous nature of Al interference within the cell, studying Al toxicity has been a largely intractable field of study. Nevertheless the research community studying Al-dependent root growth inhibition has made significant and continual progress in the pursuit to understand this complex real world agricultural problem. The primary site of Al toxic growth inhibition is at the root and more specifically at the root tip. This is expected since the primary site of contact of Al with the plant is between the soil solution and the root. The immediate effect of Al toxicity is reduced elongation of the root, which can occur as quickly as 30 minutes to two hours . While there is extensive documentation of short-term responses of the root,flower buckets wholesale upon extended treatment with Al, the root begins to swell with distorted morphological integrity, root discoloration, and differentiation of cells.
Meristematic and root cap cells also become increasingly more vacuolated and this, in part, contributes to their increase in cell volume despite inhibited ability to carry out growth by cell division. There are also changes in root cell patterning, irregular cell division, alterations in cell shape, cell wall thickening, callose accumulation, disintegration of the cytoskeleton, formation of myelin figures, alteration of the plasma membrane, and the production of reactive oxygen species . The rapid inhibition of root growth is likely caused by interference with cell elongation, while long-term inhibition of root growth is likely due to inhibition of cell division . While it is known that the primary effect of Al toxicity occurs at the root tip, the response to Al at the tip can vary. At the apex of the root tip is the root cap, which protects the apical meristem of the root and the meristematic zone. It is comprised of a pool of border cells that excrete lubricating saccharides and these cells are continually, acropetally produced and sloughed off the root tip from the friction of the growing root through the soil. The meristematic zone is the zone of cell division, which contains the root apical meristem . Within this region there is a core of slow dividing cells called the quiescent center , which produces the rapidly dividing initial cells . Above this region is the zone of elongation where the root cells elongate and then begin to differentiate . Above this zone is the final developmental zone of root development called the zone of cell differentiation, where cell fates reach maturity and the root hairs develop . A study using fluorescent lifetime imaging in plants quantified Al uptake in vivo . This report confirmed that the primary sites for Al entry are the meristematic zone and distal transition zones, while Al uptake is limited in the cortex and epidermis of the mature root zone .
Al is taken up by the roots in the form of a trivalent cation and can bind to a vast array of biochemical sites once internalized because it has a higher affinity for anionic targets in comparison to ions such as Mg2+ and Ca2+. Uptake of Al into cells is hypothesized to occur via ion channels, particularly those that transport cations of similar radius like Fe2+, Mg2+ or Ca2+ , and is proposed to enter the symplast via an Nramp-like transporter, Nrat1 . Al accumulation within root tissue can be rapid upon exposure. In soybean roots, Al can accumulate in the symplasm of the three outer cortical cell layers after only 30 minutes of treatment . Rapid uptake is also observed in Arabidopsis roots, where significant levels of Al accumulate within 1 hour of exposure . This accumulation of Al is not only very rapid but also disrupts cellular activities and induces stress signaling. Determination of Al-induced toxicity is complicated by interference with mineral absorption at the root apex, resulting in nutrient deficiency symptoms in plants grown on Al toxic soils. This is particularly true for the divalent cations Ca2+, Mg2+, and K+ as well as phosphate of which low pH soils inhibit their absorption by roots . In addition, Al has been found to competitively inhibit uptake of Ca2+, Mg2+ and K+ and the addition of these cations to the soil can ameliorate the toxic effects of Al . Impairment of ion uptake occurs via blockage of ion channels in the plasma membrane and is likely the cause of the interruption of intracellular Ca2+ and Mg2+ homeostasis and contributes to the rapid depolarization of the plasma membrane following Al exposure .While it is hypothesized that uptake of Al into cells can occur via ion channels, an Nramp-like transporter, Nrat1 , was identified in rice capable of transporting Al into the cell . This transporter is specific for Al and does not transport any other divalent metal ions such as iron, manganese and cadmium, or any Al-organic acid complexes. Knockouts of Nrat1 are hypersensitive to Al but loss of this transporter did not affect the sensitivity to other metals, and causes reduced internalized Al and increased accumulation of Al at the cell wall.
It is hypothesized that the increased Al sensitivity is due to the absence of cellular Al detoxification mechanisms with organic acids since the plant is no longer able to transport Al into the cell and instead causes accumulation of Al at the cell wall. This mechanism of Al tolerance utilizing Nrat1 for Al internalization followed by detoxification is a more effect strategy than allowing Al to accumulate at the root cell wall because of the inhibitory effects of Al to root growth by limiting cell wall elongation . These changes, as well as an increase in cytosolic Ca2+, are required for the induction of callose synthesis, which is a hallmark Al stress response in the root apex that acts to isolate cells from intercellular transport of Al . Internalized Al has both living and non-living binding sites within plant cells. Al begins to cause its toxic effects at the apoplastic cell wall and symplastic plasma membrane of root cells. As the cell wall is a primary site of Al accumulation, interaction with the cell wall accounts for more than fourfifths of total Al uptake . This initial rapid phase of apoplastic Al accumulation by the roots is due to pectin content of the cell wall and the degree of methylation of the pectin, which is controlled by pectin methylesterase . Intact roots of maize have increased Al accumulation and Al dependent root inhibition when pectin methylesterase function was enhanced . Al treatment also causes reduced root cell wall flexibility and binding of Al to the pectic matrix may prevent cell wall extension physically and physiologically by decreasing the effectiveness of cell wall-loosening enzymes . Al accumulation in the symplasm has been a historically controversial topic, but has now been clearly established. Concentrated levels of Al have been found in organelles like endosomes and vacuoles, and also in the cytoplasm where Al disrupts microtubule and actin dynamics . Additionally, Al is capable of infiltrating and binding to any anionic sites within the cell,flower harvest buckets including the nucleus where the negatively charged phosphate backbone of DNA is a site of Al accumulation. Al rapidly alters the plasma membrane by interactions with membrane lipids and proteins, which causes changes in its structural properties such as membrane fluidity, permeability and protein-dependent transport . Al also induces membrane depolarization, specifically in the distal transition zone of the root tip, which may be related to the inhibition of the H+-ATPase activity and may lead to the disruption of H+ homeostasis in the cytosol . These changes in the plasma membrane properties caused by Al affect ion transport capabilities, such as causing a rapid decrease of K+ efflux without changing K+ influx . Al-induced damage of membrane integrity also may be related to Al- enhanced oxidative stress through the formation of reactive oxygen species leading to lipid peroxidation and protein oxidation . Oxidative stress genes have been shown to be induced upon Al treatment and over expression of these genes have conferred Al tolerance .
Even though there are such drastic changes in plasma membrane structure and function, there is no evidence that there is a requirement for a severe disruption of the plasma membrane to induce root growth inhibition . It appears that Al activates signal transduction pathways that lead to the observed symplastic disorders, with a rapid increase in cytosolic Ca2+ playing a major role in this effect . Since Al toxicity reduces the absorption of Ca2+ by the root, the source of Ca2+ is likely from the apoplast due to Al liberating bound Ca2+ to the cell wall. Al also disrupts the plasma membrane potential, which activates Ca2+ channels, allowing for a burst of Ca2+ to enter the cell; however, it cannot be ruled out that Al may cause a release of Ca2+ from symplastic Ca2+ reserves as well . Increasing Ca2+ levels within the cell due to Al exposure can partially explain why callose accumulation and also cytoskeleton disorganization occurs . Al entering the symplast accumulates within the cytoplasm and results in destabilization of the cytoskeleton. Disorganized arrangements of actin filaments in the stele cells of the transition zone of maize roots occur upon Al uptake. Gene expression of actin, as well as profilin, an actin-binding protein that regulates the polymerization of actin filaments and plays a role in cell elongation, is inhibited by Al exposure . A further effect on the cytoskeleton is the reorientation of micro-tubules and micro-filaments, which are made of actin monomers. This could explain the swollen root tip that is symptomatic of Al toxicity and associated with cell cycle arrest as well as endoreduplication that results in abnormal cell plate division and spindle formation . As a trivalent cation, Al has the capability of indiscriminately binding to a wide range of negatively charged bio-molecules within the cell including sites within the nucleus. Al can displace other cations like Ca2+, Mg2+, and K+, thus inhibiting or altering the function of the structures with which Al associates . For example, Al has been found to bind to ATP with more than 100 times the affinity of Mg2+, suggesting that Al is generally inhibitory to ATP requiring enzymatic reactions that depend on an Mg-ATP complex to function . Specific toxic effects include inhibition of Ca2+ influx across the membrane through blockage of Ca2+ channels, disruption of H+ homeostasis, neutralization of the zeta potential at the membrane surface, and inhibition of H+ flux mediated by the H+- ATPase . Al is also known to interfere with the production and transport of plant hormones. Ethylene and auxin both have been shown to synergistically affect root growth, as ethylene stimulates auxin biosynthesis and basipetal auxin transport toward the elongation zone. The interference of Al with this hormone-mediated development will cause inhibition of root cell elongation. It has been shown that there is a link between ethylene and auxin signaling in Al toxicity, indicating that there is a burst of ethylene upon Al treatment, followed by auxin biosynthesis. This suggests that ethylene serves as a signal to cause downstream changes in auxin distribution in roots by interacting with AUX1 and PIN2 proteins, leading to inhibition of root elongation in the presence of toxic Al . Nitric oxide production is also negatively effected by Al treatment, which is a signaling molecule for various responses to biotic and environmental stresses. Under normal growing conditions roots express nitric oxide at the root cap statocytes, quiescent center and the distal transition zone, but upon Al treatment, nitric oxide production is abolished in the distal transition zone only .