Let us make an in-depth study of the modern view of solute transport across membranes in plant.
The mineral salts are absorbed by plants in their ionic form and so is their transport within plants. Certain solutes such as sugars are however transported across the membranes in uncharged state.
For uncharged solutes (non-electrolytes), their movement across the membrane depends upon their concentration gradient i.e., gradient of chemical potential only on two sides of the membrane. But, in case of charged solutes or ions (electrolytes) the situation becomes different.
Since, charged solutes or ions carry an electric charge, their movement across the membrane depends not only on their chemical potential but also on their electrical potential. In other words, transport of ions across the membrane depends upon their electro-chemical potential gradient. Therefore, electrical properties of the cell or its Trans membrane potential is very important component of ions transport through membranes.
In either case, the transport of solutes across the membrane is called as passive transport if it is along the chemical potential gradient or electrochemical potential gradient (for non- electrolytes and electrolytes respectively). When solute transport across the membrane occurs against the chemical potential gradient or electrochemical potential gradient, it is called as active transport and requires additional input of energy (Fig. 7.6).
The movement of solutes into the cytosol through membrane (such as plasma membrane or tonoplast) is called as influx while their exit from the cytosol is termed as efflux. In recent years much work has been done on permeability of cell membranes especially plasma membrane and tonoplast (vacuolar membrane) and various trans membrane transporters (proteins) have been identified in them which enhance movement of solutes across such membranes. These transporter proteins are highly specific with complex structure and different models have been given by scientists to explain their functioning.
These membrane transporter proteins can be grouped in three categories:
(i) Ion-channels,
(ii) Carriers and
(iii) Pumps
1. Ion-Channels:
i. Ion-channels are trans-membrane proteins which function as selective pores through which ions can diffuse easily across the membrane.
ii. Ion-channels are usually highly specific for one or limited number of ion species. The specificity depends upon the size of the pore and density of surface electric charges on its interior lining more than on selective binding of ions.
iii. Transport of ions through channels is always passive.
iv. The channels are not open all the time but are ‘gated’ (Fig. 7.7). The gates open or close in response to external stimuli that include, (i) voltage changes, (ii) light, (iii) hormone binding and (iv) ions themselves. When gates are open, the ions can diffuse through the channels but not when they are closed.
The channel proteins are believed to contain a sensing region or sensor which responds to the appropriate stimulus by changing conformation of channel protein opening the gate.
v. Because the ions carry a charge and are mobile, their diffusion across the membrane channel establishes an electric current, which can be detected by a special technique which is known as ‘patch-clamp electrophysiology’.
vi. Patch-clamp studies have shown that for a given ion such as K+, a given membrane has variety of channels which may open in different voltage ranges or in response to different stimuli such as K+ and Ca2+ concentrations, pH, protein kinases and phosphatases etc.
vii. Ions can diffuse through an open channel with rapidity as high as 108 s-1.
viii. Those channels which allow inward transport of ions (i.e., towards cytosol side), are called as inward rectifying or inward channels and those which allows outward diffusion (i.e., from cytosol to other side) are called as outward rectifying or outward channels.
ix. Ca2+ channels are inward rectifying while anion channels are always outward, (for transport of such ions in reverse direction, active transport mechanisms are required)
x. K+ is exceptional. It can diffuse inward or outward across the membrane through channels depending upon more negative or more positive membrane potential respectively.
xi. Many channel proteins are inducible i.e., they are synthesized by the cell when a particular solute is available for absorption.
(Some slow vacuolar (SV) channels may be present on tonoplast which allow diffusion of some cations and also anions from vacuole to cytosol).
2. Carriers:
i. These trans-membrane transporter proteins do not form pores in membrane, instead they selectively bind the solute to be transported to a specific site on them. This causes conformational change in carrier protein which exposes the solute to other side of the membrane. After the solute is released from the binding site, the carrier protein reverts back to its original conformation to pick up a fresh solute molecule or ion (Fig. 7.2). Thus, binding and release of solute through carrier is similar to an enzyme catalysed reaction.
ii. Carrier mediated transport of solutes enables transport of much wider range of solutes, but is slower (about 104 – 105 s-1) as compared to channel mediated solute transport.
iii. Carrier mediated solute transport may be of two types:
(i) Passive transport and
(ii) Active transport.
Passive Transport:
Carrier mediated passive transport of solutes occurs along the electrochemical potential gradient and does not require expenditure of energy. This has also been called as facilitated diffusion. According to some scientists, both carrier mediated passive transport and channel mediated transport should come under the purview of facilitated diffusion.
Active Transport:
Carrier mediated active transport of solutes takes place against the electrochemical potential gradient and requires additional input of energy that chiefly comes from hydrolysis of ATP. In such cases, the carrier proteins are called as ‘pumps’ and the transport of solutes is called as primary active transport because it directly utilizes energy from hydrolysis of ATP.
3. Pumps:
As mentioned earlier, the membrane transporter proteins involved in primary active transport of solute are called as pumps. Most of the pumps transport ions such as H+ and Ca2+ across the membrane and are known as ion-pumps. Some pumps (such as those of ABC transporters category) may also transport large organic solutes across the membranes.
Ion-pumps may be of two types:
(i) Electro neutral pumps and
(ii) Electro genic pumps.
Electro neutral pumps are those which are associated with transport of ions with no net movement of charge across the membrane. For example H+/K+-ATPase of some animal cells, pumps out one H+ for each K+ taken in with no net movement of charge. Therefore, it is an electro neutral pump.
Electro genic pumps on the other hand, transport ions involving net movement of charge across the membrane. For example, H+-ATPase found in plant and animal cells, pumps out H+ with net movement of one positive charge. Therefore, it is an electro genic pump. (The Na+/K+ – ATPase of animal cells such as neurons, is also an example of electro genic pumps because it expels three Na+ ions for every two K+ ions taken in resulting in net outward movement of one positive charge.)
H+-ATPase, H+-PPase and Ca2+-ATPase are most common electro genic pumps in plant cells and their direction is outward. (Therefore, other mechanisms (secondary active transport) are required for uptake of most of the mineral nutrients).
A brief account of some of the most common pumps in plant cells is as follows:
(i) Proton-ATPase Pumps (H+-ATPases):
These pumps are also known as P-type ATPases and are found in plasma membrane, tonoplast and possibly other cell membranes. These are structurally distinct and operate in reverse of F-type ATPases i.e., they hydrolyse ATP instead of synthesizing it (ATPases of mitochondria and chloroplast are also known as F-type ATPases)
Fig 7.8 shows a model of plasma membrane H+– ATPase (also known as P-type ATPase). This enzyme protein is a single chain polypeptide with 10 hydrophobic trans membrane segments or domains (only three of these are shown as helical coils while others are shown as cylinders in the figure).
These segments are joined by hydrophilic loops which project in cytosol and cell wall (apoplast). The ATP binding site is believed to be an aspartic acid residue (D) situated on loop connecting 4th and 5th segments towards cytosilic side. Hydrolysis of ATP causes conformational change in the protein and one H+ ion is transported from cytosol to outside across the plasma membrane.
The H+ – ATPases of plasma membrane and tonoplast are different.
Plasma membrane H+-ATPases:
i. These are characteristically inhibited by vandate ions (VO3–) but are insensitive to other ions such as NO3–.
ii. Single proton (H+) is trans located for each molecule of ATP hydrolysed.
Vacuolar H+-ATPases:
i. These are insensitive to VO3– but are strongly inhibited by NO3–.
ii. Two protons (2H+) are trans located for each molecule of ATP hydrolysed.
iii. Resemble structurally to F-type ATPases of mitochondria and can be separated into two complexes analogous to F0 and F1, out unlike the former are not inhibited by oligomycin or azides.
(ii) Proton-pyrophosphatases (H+– PPases):
i. There are mainly found in tonoplasts but may also occur in membrane of Golgi-bodies. They pump protons into the lumen of vacuole and Golgi-cisternae.
ii. These pumps appear to work in parallel with vacuolar ATPases to create protons gradient across the tonoplast. This enzyme protein consists of a single polypeptide chain with molecular mass of 80 kD and utilizes energy from hydrolysis of inorganic pyrophosphate (PPi).
iii. Free energy released by hydrolysis of PPi is less than that obtained from hydrolysis of ATP. Vacuolar H+ – PPase transport only one H+ per PPi molecule hydrolysed.
(iii) Calcium Pumping ATPases (Ca2+-ATPases):
i. These are found in plasma membrane, tonoplast and possibly other cell membranes such as those of chloroplasts and ER.
ii. These pumps couple hydrolysis of ATP with translocation of Ca2+ across the membrane.
(iv) ATP-Binding Casette Transporters (ABC Transporters):
Certain large metabolites such as anthocyanin’s and other secondary plant products are removed from the cytosol and transported across the tonoplast to the vacuole through ABC- transporters located on tonoplast that consume ATP directly. Recently ABC transporters have also been reported from plasma membrane and also mitochondria.
Secondary Active Transport—Symport and Antiport:
A large number of nutrients are transported across the cell membranes against their chemical potential or electro chemical potential gradients by secondary active transport mechanism that does not utilize energy liberated by hydrolysis of ATP directly but indirectly through the energy stored in proton-electrochemical potential gradient across the membrane or proton motive force.
The electro genic proton-ATPase (H+-ATPase) pumps serve as proton trans locating carrier proteins and free energy of hydrolysis of ATP is conserved in the form of proton gradient across the membrane (more protons accumulating on outer side). This proton gradient together with normal membrane potential contributes to proton electrochemical potential gradient or a proton motive force which tends to move protons back across the membrane through specific carrier proteins located elsewhere on the same membrane.
When protons return back to cytosolic side, the proton motive force generated by electro genic H+ transport can be utilized to drive transport of other solute molecules or ions against their chemical or electrochemical potential gradient through the same carrier protein through which H+ is returning back to cytosol. This is called as secondary active transport and because ions or molecules of two different substances (H+ and other solute) are being transported at the same time through the some carrier protein, this process of solute transport is also called as cotransport mechanism.
Secondary active transport or cotransport mechanism is of two types:
(i) Symport and
(ii) Antiport.
Symport:
When influx of protons is coupled with movement of other solute in the same direction, the cotransport mechanism is celled as symport and the carrier protein is called as symporter (Fig. 7.9A).
Antiport:
When influx of protons is coupled with efflux of other solute, the cotransport mechanism is called as antiport and the carrier protein involved is called as antiporter (Fig. 7.9 B).
Fig. 7.10 gives an overview of the various solute transport processes through plasma membrane and tonoplast in plants.
