Growth and development of plant are dependent on photosynthesis and transpiration. These two processes are controlled by regulation of water movement and homeostasis. As universal solvent, water is undeniably crucial in transport of solutes and minerals throughout a whole plant system. However, excess of water in the plant's system will pose as much problem if there is water deficient hence the need of a tight regulation of water balance in a plant.
In plants, a long distance transport involves water transport through the vascular tissue i.e xylem and phloem. By means of bulk flow, water transport from root to shoot for example, almost always do not encounter membrane barriers. This type of long distance transport usually employs the apoplastic pathway in which water molecules move across cell wall continuum. For instance, xylem sap which moves along the vascular bundle has no direct association with aquaporins. Another type of transport is a short distance and non-vascular tissues transport which requires movement of water molecules and/or solutes across biological membranes. Transmembrane water transport occurs both by diffusion through lipid bilayer of plasma membrane and across water channel, aquaporins. Cell-to-cell transport can either take up a symplastic pathway which transports water in cytoplasmic continuum of adjacent cells through plasmodesmata, or by transcellular pathway of which water molecules are transported across plasma and vacuolar membrane.
2.1 Aquaporins
(Zhao et al., 2008) defined aquaporins as channel-forming membrane proteins which are capable to combine high flux with high specificity for water across biological membranes. This description alone however is insufficient in terms of its function as aquaporins proved to permeate non-polar solutes such as urea and glycerol, polar ammonia gas (NH3), non-polar carbon dioxide gas (CO2), and reactive oxygen species (Kaldenhoff and Fischer, 2006); (Tyerman et al., 2002); (Johansson et al., 2000, Heinen et al., 2009)).
Figure 1: Topology of trans-membrane aquaporin protein. The protein consists of six trans-membrane alpha helices protein (not depicted as alpha-helices here) numbered I to VI accordingly. Five loops (A-E) adjoined the six proteins together. Loops B and E contained conserved NPA (asparagine-proline-alanine) motifs, forming short α helices that fold back into the membrane from opposite sides. C (carboxyl terminus) and N (amino terminus) face the cytosol (Kruse et al., 2006).
Aquaporins belong to a major intrinsic protein (MIP) superfamily. This water-channel was first discovered from human erythrocytes named AQP1 and thought to be common in almost all living organisms. The topology of aquaporins is illustrated in Figure 1 above. Intracellular loop B and extracellular loop E are of particular interest as these two loops contain highly- conserved NPA (asparagine-proline-alanine) motifs (Kaldenhoff and Fischer, 2006); (Tyerman et al., 2002)). 180 degree orientations of the two NPA motifs create an aqueous pathway through the proteinaceous pore (Kruse et al., 2006). It is widely accepted that all aquaporin-like proteins assembled into tetramers though each monomer in the tetrameric conformation is a functionally independent water channel (Kaldenhoff et al., 2008))
Usually, aquaporins of higher plants are categorized into four subclasses, most likely determined by their distinct subcellular localizations. Tonoplast intrinsic proteins (TIPs) and plasma membrane intrinsic proteins (PIPs) are expressed predominantly in vacuolar membrane and plasma membrane respectively. Small intrinsic proteins (SIPs) are named accordingly due to its short cytosolic N-terminal region compared to other plant MIPs, and mostly located in the membrane of endoplasmic reticulum. Nodulin-26 like intrinsic proteins (NIPs) were initially discovered in peribacteroid membrane of nitrogen-fixing nodules of leguminous plants and later were also found localized in plasma membrane of other plant species. The classification of the aquaporins based on the subcellular localization has been recently questioned in (Maurel et al.)(2009) as more aquaporins-like proteins have been discovered elsewhere than their predominant site of localisation. New subclass of MIPs called X intrinsic proteins (XIPs) has been discovered in moss Physcomitrella patens, and in tomato and poplar (Maurel et al., 2009).
2.2 Root to xylem stele
Water moves from root to xylem stele by a radial transport but incorporates two different water pathways i.e apoplast and symplast pathways. (Steudle, 1994) proposed root cell protoplasts considered holding a hydraulic resistance much greater than root apoplasts, hence water moves around the protoplasts. In other words, water follows the apoplastic route in preference to the symplast route when crossing the epidermis layer in the root. However, upon crossing the endodermis layer, highly suberized waterproof band called Casparian strip prevent water passage by the apoplastic route. Water molecules are forced to enter the symplast route, crossing the plasma membrane of the endodermis through the plasmodesmata and into the xylem stele. In this case, aquaporins facilitate the water transport across the endodermal cells by providing a much less hydraulic resistance in relative to the apoplast route. This is crucial especially during high rate of transpiration where water needs to be transported from the root towards the leaves to compensate water loss. Many aquaporins are present in the root endodermis (Kaldenhoff et al., 2008) and xylem parenchyma cells (Maurel, 1997).
2.3 Xylem transport
The axial transport of water molecules along the vascular bundle may not involve any aquaporins as the movement of xylem sap is by passing through a cell wall continuum. The vascular bundle is nearly all made of non-living cells and thus, the apoplastic route is the predominant pathway[X SURE]. However, symplastic cells encircle the xylem bundle may greatly play a part in the xylem axial transport as well as plant water transport as a whole.
In the bundle sheath, water transport is challenged with apoplastic barrier similarly to the root endodermis because sometimes the cell walls of the vascular bundles are lignified or suberized. Again, water is forced to enter symplast pathway and presence of aquaporins greatly contribute to water movement. Highly expressed of several aquaporins in the bundle sheath cells and xylem parenchyma cells such as ZmPIP1, ZmPIP2 and ZmTIP1 in maize may imply the role of aquaporins in water transport across the xylem vessels[reference mana?].
Hydraulic conductance is governed by the ratio of water flow rate to driving force of the flow. Often, the driving force is determined by the differences in water potential due to pressure (hydraulic gradient) and solute concentration (osmotic gradient) (Hachez et al., 2008). Cavitation reduces xylem hydraulic conductivity, affecting the transport of water from the root to leaf.. Stomata are forced to close in order to prevent further cavitation and leaf dehydration, which greatly distress the rate of CO2 uptake by the leaf. Normally cavitations are triggered by water stress and freeze-thaw cycle. Gas or vapour bubble formed keeps expanding, filling the cavitated vessel as a result of water molecules being pulled from surrounding tissue into the cavitation(??). One of several mechanisms to restore xylem hydraulic conductivity during embolism is proposed by (Sperry et al., 2003) which is water reloading by xylem parenchyma cells. Adjacent vessels are thought to supply the water for refilling of which solutes of sufficient sizes are secreted by the parenchyma cells into the embolized-vessel. Water molecules then follow, moving down the water potential gradient possibly mediated by aquaporins (De Boer and Volkov, 2003). High expression of PIP2 aquaporins in vessel-associated cells supports the hypothesis of water refilling during embolism is demonstrated by (Sakr et al., 2003).
Water balance in plants is partly controlled by stomatal closure in the leaf. As to allow gaseous exchange in the leaf and consequently permitting water loss during photosynthesis, stomata opening and closure is critical in maintaining between the two circumstances. Guard cell flanking the stoma is the key in stoma aperture, immediately controlled by change of turgor. Basically, accumulation of potassium ions (K+) in the guard cells from neighbouring epidermal cells lowers the osmotic potential in the guard cells. Water follows into the guard cells, increasing the volume and turgor pressure. Swelling of the guard cells causes the stoma to open due to its thin and flexible walls. By similar mechanism, stomatal closure occurs when the osmotic potential in the guard cells drop as water enters neighbouring epidermal cells. Influx and efflux of water is highly likely to be mediated by aquaporins. Accumulation of SunTIP7 and SunTIP20 mRNA in the guard cells of sunflower leaves (Sarda et al., 1997) are thought to be exclusive in the guard cells, which supports the presence and role of aquaporins in stomatal movement. In (Sun et al., 2001), bbaq1 is expressed in guard cells of Vicia Faba and none were detected in the epidermal cells. He insisted that aquaporins are exclusive in guard cells in the basis of results by (Kaldenhoff et al., 1995) and (Sarda et al., 1997) though Feng et al (2002) detected localization of RD28 in epidermis and guard-cell protoplasts of Vicia Faba. Aquaporins may also be expressed in surrounding subsidiary cells as well as epidermal cells as suggested in (Heinen et al., 2009). Various form of aquaporin is localized in different parts of the plants suggest that this water-channel is multi-functional and its role is yet to be established.
Phloem unloading of sucrose is of great importance in a plant as photoassimilates produced in the leaves are transported to plant sinks (i.e roots, growing cells). A common view is that photoassimilates proceeds along the phloem by means of diffusion through mesophyll symplasm via plasmodesmata towards the sieve elements and companion cells. Holbrook and Zwieniecki pointed out that the phloem sap unloading may also be dependent on pressure gradient, which implies a more specific and regulated mechanism is involved. Phloem unloading are categorised to 3 different stages (i)collection; photoassimilates loaded into sieve elements(SE)/companion cell(CC) of leaf (ii)transport; photoassimilates transported from leaf minor veins into sieve tubes and (iii)release; unloading of photoassimilates from SE into growing or storage cells (Holbrook & Zwieniecki). They proposed that transmembrane proteins exert control on turgor pressure and the rate of solute release from SE must related to rate of water loss in some way. Level of solute concentration in parenchyma cells adjacent to SE must be kept high so as to maintain steep solute gradient through the symplast.
2.4 Aquaporins and leaf movements
In a general sense, plant movements can be divided into often reversible turgor movements and irreversible growth movements. Many works have related leaf movements with specialized motor organs called pulvini, consisted of two functionally distinct tissues: adaxial flexor and abaxial half referred to as extensor. Reversible shrinkage of the extensor and swelling of the flexor cells is driven by changes in osmotic water fluxes, caused by changes in solute content and ion composition, mainly potassium. Shrinkage and swelling of the extensor and flexor cells induces the opening of the pulvinus. By the same mechanism, the pulvinus will close (Uehlein and Kaldenhoff, 2008). Aquaporins of PIP family was found in both flexor and extensor tissues of Samanea saman (Moshelion et al., 2002). In oppose to nyctinastic movements, tobacco leafs are found to employ an epinastic way of movements, that is movements of the leaf without the presence of specialized motor organs. Controlled by circadian and diurnal factors, NtAQP1 proves its importance in leaf movements of tobacco leaf. Expression of NtAQP1 in the leaf petioles of tobacco plants correlates with diurnal and circadian oscillation. (Siefritz et al., 2004) showed that protoplasts from leaf petioles exhibited high cellular water permeability in the morning (when leaf unfolds) and low in the evening. Transgenic tobacco plants with an impaired expression of NtAQP1 showed a reduced diurnal epinastic leaf movement due to considerably low cellular water permeability in relative to the controls. Both processes have proven the relationship between aquaporins and leaf movements but the molecular mechanism of this water-channel is yet to be elucidated.
2.5 Other role of aquaporins
It is worth to note that aquaporins also contribute in carbon dioxide (CO2) conductance in a plant. (Flexas et al.) (2006) established the involvement of tobacco aquaporin NtAQP1 in mesophyll conductance to CO2 in vivo. When aquaporins isolated from barley HvPIP2 were overexpressed in transgenic rice plants, there was an increase of internal CO2 conductance and rate of CO2 assimilation in the leaf (Hanba et al., 2004). In tandem with a high internal CO2 conductance is an increase in stomatal CO2 conductance which consequently brings about more water loss from the leaves. Hence, regulation of CO2 conductance in the leaf by the aquaporins not only influences rate of photosynthesis but also rate of water loss from the leaf, and ultimately the plant water balance.
Stress-induced aquaporins
Plants are subject to stress conditions such as drought, salinity or freeze which is partly due to their immobile characteristics. Land plants have evolved several specific mechanisms to encounter these stresses at both molecular and cellular level. Aquaporins are thought to facilitate in maintaining homeostasis under water-stress conditions (Tyerman et al 2002). However, different findings regarding the role of aquaporins have been documented between drought-adapted and non-adapted varieties of plants as well as cultivars (Lian et al., 2004). When soil dries up, hydraulic conductivity (Lp) of root usually decreases as shown in Agave deserti plants which may partly due to collapse of cortical cells, increased of suberization and embolism in xylem vessels (Vandeleur et al., 2009). Roots also shrink during drying conditions, reducing contact between soil and roots. Transgenic tobacco plants were found to wilt faster when water was limited due to overexpression of PIP1 (Aharon et al., 2003). On the contrary, a reduced resistance to water stress was observed in antisense tobacco plants with reduced expression of NtAQP1 (Siefritz et al., 2002).
Strong correlation between cellular water permeability and root hydraulic conductivity was established in (Siefritz et al., 2002) of which it was discovered that reduced expression of NtAQP1 and NtPIP1 in tobacco plants results in decrease of overall specific water conductivity of the root system. For tobacco plants that were grown in well-watered condition, PIP1 did not seem very significant in terms of water uptake and management (Siefritz et al., 2002). PIP2 was found to be more efficient as water channels than PIP1, as documented in Kaldenhoff et al. (2006) and (Zhao et al., 2008). However, when the antisense tobacco plants were grown in low soil water potential, a rise in hydraulic resistance was observed which is likely due to lack of NtAQP1. The antisense plants were not able to maintain turgor pressure above its wilting point, appearing much more drought-sensitive (Siefritz et al., 2002). So, as water becomes a limiting factor, PIP1 aquaporins significantly enhances root hydraulic conductivity and cellular water permeability, and relieve osmotic pumps to survive the plants during dry/drought periods. Vandeluer et al. (2008) also supports the hypothesis of reduction in root hydraulic conductivity in response to water stress. Two cultivars of grapevines were subjected to water stress, anisohydric Chardonnay and isohydric Grenache, where both cultivars show contrasting responses. Up-regulation of aquaporins in Chardonnay supports the importance of PIP1 in tolerance to water stress.
The more drought-tolerant Grenache, however, maintained its transcript level of VvPIP1 and VvPIP2. The failure of mercuric chloride to further reduce hydraulic conductivity in the roots led to the idea of decreasing hydraulic conductivity induces the closure of aquaporins gates (Martre et al., 2001). Hypothetically, closing of aquaporins gates will prevent water loss from the root to the soil, which has a relatively lower water potential than the roots. Desert plants and aspen seedlings are thought to employ this strategy of reducing the expression of aquaporin to prevent water loss to the soil. Desert plants exhibited a significant increase in Lo after rewatered which may be explained by its adaptation to environmental condition with limited rainfall (Vandeluer et al., 2008) i.e desert plants are more drought-resistant. This suggests the roles of aquaporins are entirely different depending on types of plants and their environment.
Plant aquaporin conductivity was suggested to be regulated by gating mechanisms involving protein phosphorylation under water-stress conditions and protonation after cytosolic acidification after flooding (Fischer and Kaldenhoff, 2008). Simultaneous closure of all Arabidopsis PIPs upon anoxia (absence or deficiency of oxygen in tissue) was reported due to protonation of histidine in loop D (Tornroth-Horsefield et al., 2006). In the same work, it was also discovered that closure of SoPIP2:1 of spinach is triggered by dephosphorylation of two serine residues in the cytosolic loop B and the carboxy-terminal region.
Figure 2: Structural mechanisms of aquaporins gating in plasma membrane aquaporins (PIPs) in different conditions. In normal condition, aquaporin proteins are phosphorylated (indicated by P) and opens the gate. During drought stress, the gate closes due to dephosphorylation of two serine residues (not depicted here). During flooding too, the gate closes, due to protonation (indicated by H+) of a histidine residue in loop D of the aquaporin-like protein (Tornroth-Horsefield et al., 2006) .
Conclusion
Water constitutes as much as 95% of a plants' weight, implying of great importance water is in a plant system. Presence of aquaporins is undoubtedly crucial to fine-tuned water regulation especially in higher plants where a higher degree of compartmentation of plant cells is involved (Johannson et al. 2001).
