A hypothesis concerning callose

ABSTRACT

Background Concept: Certain proteins and the glucose monomer have spacing of their carbonyl oxygen atoms that match the spacing of the oxygen atoms of hexagonal ice. This opens the possibility that a sequence of linked glucose residues may have a sequence of equally spaced carbonyl oxygen atoms. Hypothesis: Callose In plants is a duality consisting of the callose itself and a layer of ordered water whose oxygen atoms are hydrogen bonded to the carbonyl oxygen atoms in the callose. The atomic basis of the hypothesis is that the 1–3 linkage between glucose residues in callose results in equally spaced carbonyl oxygen atoms within and between residues. Properties of Callose/Ordered Water: The physical properties of the duality are the properties of callose itself: 1) it is immobile, 2) it can be created and dissolved, 3) it can exist at a submicrometer to micrometers space scale. The electrical properties of ordered water in a botanical platform are not known at the present time. They can be derived only from limited data in non biological platforms and inferences from the electrical properties of ice. These properties are 1) proton movement is governed by the Grotthuss mechanism, 2) there is insignificant movement of non-protonic ions and larger molecules through the ordered water, 3) proton movement is isotropic. Proposed Functionality of Callose/Ordered Water: Known locations of callose were examined theoretically to determine the functionality of a callose/ordered water duality. These locations were sieve plate pores, plasmodesmata and pollen tubes, stomatal guard cells, companion cell/sieve tube complex and micro and megasporocytes. Protonic Circuits: In a botanical context, protonic circuits at a single cell and supracellular level take the form of a proton microloop wherein callose/ordered water is one component in the loop. These circuits use both the enhanced proton mobility and the ion blocking ability of ordered water.

Background to the hypothesis

Callose presence in plants

Callose is ubiquitous in algae and land plants.^1–4 It has a transient presence and/or a permanent presence. Examples of non transient locations are the sieve plates connecting the lumen of adjacent sieve cells in the phloem and the plasmodesmata between the lumen of the sieve cell and the companion cell. Callose is present in developing pollen tubes.⁵ Callose is present in the septae of algae during cytokinesis.⁴ It can be established very rapidly in the case of mechanical damage to sieve tubes to block passage. Such callose is termed wound callose.⁶ In spite of its widespread presence, the function of callose is still unsettled. The common interpretation is to attribute the functionality of callose to a plugging agent to block passage of pathogens through the sieve pores in case of injury to the plant. This does not explain its presence in stomates or in pollen tubes. The purpose of this paper is to explain that the diverse presence of callose is derived from the functionality of ordered water.

Hydrogen bonding between water in crystalline form and biological compounds

Warner observed that oxygen atoms in proteins and other biological compounds can be hydrogen bonded to oxygen atoms in water in crystalline form.⁷ The requirement for this bonding is a 0.48 nanometer spacing between the bonds. Franks extended Warner’s observation to include sugars, steroids and triglycerides.^8,9 Table 2 of Franks’ paper illustrates the precise location of the bonds between hexagonal ice and some biologically important reactants including glucose.

Table 2.

Mobilities of Selective ions¹⁰

Particle	Mobility, meter squared/sec volt
Ion (K+ in water)	~ 5 * 10E-8
Proton in water	3*10E-7
Ion (e.g. LI+) in ice	<< 1 * 10E-12
Proton in ice	1* 10E-5 to 1* 10E-4

^{10 Table 5.2}

Proton mobility in liquid water and ice

Bockris and Reddy extensively discuss proton movement in water in liquid form and proton movement in water in crystalline form.¹⁰ The fundamental parameter describing proton movement is mobility. This is the proportionality constant relating the electrical potential gradient to the velocity of proton movement. They point out the strong differences in mobility of proton movement in liquid water and ice.¹⁰ Proton movement in ice is accomplished by a special mechanism wherein an individual proton does not move through the crystal, rather an exchange of protons (or hydronium ions) occurs from one oxygen to another. The net result is a movement of a protons through the crystal, but the same proton that enters the crystal at one end is not the proton that leaves the crystal at the other end.

This is not a new mechanism. Grotthuss described a “proton jumping” mechanism in 1806.^11,12 The basic requirement is the presence of ordered water. Ordered water is liquid water that has its oxygen atoms aligned in a constant, fixed spacing. The liquid water becomes crystalline or ice-like.

The hypothesis

Callose exists in algae and plants as a duality consisting of the callose itself and a layer of ordered water hydrogen bonded to the callose.

Mechanism of formation of the duality of callose and ordered water

The shape of the callose polymer in unembedded form may well be helical.¹³ But callose normally is present embedded in a substrate. The Author makes the assumption that the form of the substrate forms the shape of callose under normal non-injury related conditions. At a small space scale of submicrometers the shape of callose becomes linear. The discussion below begins with this linear form of the polymer.

The mechanism of formation of the duality will be discussed in three steps. Figure 1(a) shows the atomic arrangement of three repeating glucose residues in a 1 −3 linkage.¹⁴ The repeating unit forming the chain of residues is the middle residue in this figure.

Figure 1.

(a) Molecular structure of callose showing three repeating glucose residues and 1–3 linkage between residues. Structure taken from Salisbury and Ross, 1992, Fig. 8–19 Distance between #2 and #4 carbons within a residue taken from Warner, 1968, Table II. (b) Distance between residues. The linkage between the #3 carbon of one residue and the #1 carbon of the adjacent residue results in a distance of 0.48 nm between the #4 carbon of one residue and the #2 carbon of the adjacent residue (shown in red). The result is a sequence of carbonyl oxygen atoms spaced 0.48 nm apart. (c) Carbonyl oxygen atoms hydrogen bonded to water oxygen atoms. The water is thereby “ordered”. Brown ovals: carbon atoms; yellow ovals: linkage oxygen atoms; orange ovals: ring oxygen atoms; dark blue ovals: carbonyl oxygen atoms; light blue ovals: water oxygen atoms; black ovals: hydrogen atoms. Dashed lines are hydrogen bonds.

Franks pointed out a distance of 0.48 nm between the 1 and 3 carbon atoms of the glucose monomer matched the hexagonal spacing of oxygen atoms in ice.⁸ The glucose monomer exhibits a symmetry between the #1 and #3 carbons and the #2 and #4 carbons. ¹⁵ The author then assumed that the distance of 0.48 nm holds for the #1 and #3 carbons and the #2 and #4 carbons.

Figure 1(b). illustrates the next step. The 1–3 linkage pulls the two residues closer together such that the #4 carbon atom on one residue is 0.48 nm from the #2 carbon atom on the adjacent residue. This results in an equal spacing of the #2 and #4 carbon atoms over the chain of residues. This is the atomic level basis of the hypothesis.

Figure 1(c) illustrates the result of this equal spacing. The water molecules at the surface of the callose align such that there is hydrogen bonding between the carbonyl oxygen atoms on the glucose residues with the water oxygen atoms of the water. This bonding forms a dual rigid atomic structure consisting of the glucose residues in the callose with the water molecules in the water. In this sense, if there is water at the surface of the callose, you can’t get callose without ordering the water at the surface of the callose.

The height above the callose wherein this orderliness exists is not known. Some insight into this dimension is possible from the macroscopic observation of the water holding capacity of callose.¹⁶

The diagram in Figure 1(c) illustrates only a single set of bonds in the dimension extending into the page. Presumably this same orderedness exists in strands of callose and water in this second dimension.

Characteristics of ordered water

The characteristics of the callose/ordered water duality can be divided into physical characteristics and electrical characteristics.

Physical characteristics of callose/ordered water

The physical characteristic are based on the presence of callose itself. This is widely documented.^3,17 The focus of these characteristics is the ability of callose to seal a passage in a plant. This is the wound callose or definitive callose.^{3 p364, 6 p196, 18 p514}.

In summary, the physical characteristics of callose/ordered water are: 1) it is immobile, 2) it can be created and dissolved and 3) it exists to a physical extent in the order of sub micrometers to micrometers. These characteristics are no more than descriptions of the physical characteristics of callose itself up to the present time.¹

Electrical characteristics of callose/ordered water

The electrical characteristics of callose/ordered water center around the electrical characteristics of ordered water. Ordered water is crystalline liquid water. But crystalline solid water or ice is well known. The description of the electrical characteristics of ordered water can be viewed as lying somewhere between the electrical characteristics of unordered water and ice. Figure 2(a–c) describes this method of analysis of the electrical characteristics of ordered water. One considers three categories of water: 1) unordered water, that is, liquid water with little or no crystalline structure, 2) ordered water, that is, a crystalline form of liquid water, 3) ice, that is, solid crystalline water.

Figure 2.

(a) Cross section of a volume of unordered water. The volume has no crystllaine properties. This represents the other limiting condition of ordered water. This leads to the entries in Table 2. (b) Cross Section of the Duality of a layer of callose oxygen atoms (dark blue ovals) and layers of water oxygen atoms (light blue ovals) bonded to the carbonyl oxygen atoms. Hydrogen atoms not shown. The ordered water is crystalline and as such approaches the mobility of ice. The red oval indiates an oxygen atom which is in the process of transferring a proton. (c) Cross Section of a volume of ice showing the patterm of oxygen atoms. Movement of protons would occur via transfer from one oxygen atoms to the next¹⁰, Fig. 5.11. Ice is crystalline and as such all the oxygen atoms are potential transfer agents. This leads to the entries in Table 1. Salts and ions have been expelled from the crystal upon freezing (in sea ice) (Lars Chresten Lund-Hansen, personal communication).

In Category 1, that is, unordered water, ion movement is described in terms of mobility, molar conductivity and equivalent conductivity.¹⁰

Table1 gives the vales of molar conductivity of common ions

Table 1.

Molar Conductivity of Some Common Ions^{10, 19}

Ion Type	Molar Conductivity at Infinite Dilution, (siemens meter squared/mole) * 10−4
H+, (Proton)	350
K+	73
Cl−	76
NO3−	71
Li+	39
½ PO43-	69

10 Molar conductivity is defined by Eqn. 4.133, p368. 19 Section 5.111

In Category 3, that is, ice, ions are expelled during formation so the molar and equivalent descriptions of conductivity are not applicable (Lars Chresten Lund Hansen, personal communication). Protons occupy a special status in ice because they move through ice via the Grotthuss mechanism. Table 2 describes the mobility characteristics of protons and common ions

The mobility of protons in ice is two orders of magnitude greater than the mobility of protons in unordered water.

In Category 2, that is, ordered water, there is only limited data describing the values of conductivity and mobility. Taylor gives the conductivity value of ordered water as 3.5*10E−3 S/m.²⁰ Schultz states that salt is insoluble in ordered water.²¹ This indicates that not only is proton mobility enhanced in ordered water, the mobility of other ions is strongly inhibited in ordered water. Presumably, if ions under an electrical potential gradient are not able to move through ordered water, larger particles such as pathogens would have an even more difficult passage.

Based on this limited data, callose/ordered water has three useful electrical characteristics: 1) Proton movement through ordered water is isotropic, 2) Proton movement through the ordered water is strongly enhanced compared to unordered water, 3) Movement of other ions and larger entities is strongly inhibited. The isotropic characteristic is simply the affirmation that protons in ordered water move the same from left to right as from right to left. The degree of other ion inhibition within the ordered water can only be inferred at the present time. Ball visualizes ordered water as a “protonic wire” because of the enhanced movement of protons.¹²

It is possible to back calculate from the reported value of conductivity and determine the number of oxygen atoms actually engaged in transferring protons at any one time in a cross section of ordered water. Each oxygen in a cross section of ordered water is a potential carrier of a proton through the Grotthuss mechanism. This means that conductivity is measured by the number of potential paths or oxygen atoms in a cross section. Assuming the spacing of oxygens in ordered water is the same as ice, that is, 2. 76 * 10E−10 between atoms, the number of oxygen atoms in a distance of one meter in both the x and y directions is 1.3*10E19. If the atom transfers a proton and the proton carries a charge of 1.6*10E−19, then the charge crossing a one meter by one meter cross section in one second is 2.1 coulombs. This leads to a current of 2.1 amperes. At an imposed field of one volt per meter, the conductivity value measured by Taylor yields a value of coulombs in one second of 3.5*10E−3.²⁰ The ratio of oxygen atoms engaged in the Grotthuss mechanism in a cross section is 2.1/3.5*E10-3 or 600. This indicates that at any one time in a cross section of ordered water, one oxygen atom in 600 is engaged in the process of transferring a proton under a field of one volt per meter. This computation illustrates the path analysis required in analyzing proton transfer with the Grotthuss mechanism.

Testing the hypothesis

In vitro methods

The carbonyl oxygen spacing of callose itself illustrated in Figure 1(c) can be checked.

The duality can be tested in vitro in a number of ways. Callose can be laid down on a surface. The water contiguous to the surface can be tested for immobility. A test of this type can also determine the extent of the orderedness of water in the dimension above the layer of callose. This type of test (although not specifically related to ordered water) has already been performed by Eschrich in examining the water holding capacity of callose.¹⁶ Callose can also be laid down on a surface and the presence of ordered water can be tested by measuring the mobility of protons within a known volume of water above the surface under an electrical potential gradient. A test of this type can also determine the level and physical extent of the enhanced mobility.

The index of refraction of ice (1.31) differs from liquid water (1.0). The presence and crystallinity of ordered water may be testable by measuring differences in the index of refraction. This may be possible at a microscopic space scale. Reflection of light may also be a possibility.

Conductivity of protons and other ions can be used as a measure of the extent of ordered water above a surface of callose.

A fourth test of the hypothesis can be to determine the penetration of ions other than protons through the ordered water under an electrical potential gradient

More complex methods of analyzing ordered water have been employed for industrial purposes.^21,22

In vivo methods

The in vivo method involves testing for the presence of ordered water. Callose deposited on a cell wall in tissue at the surface of a plant would be ideal. Stomatal guard cells are a possibility. The method employed by Weisenseel in pollen tube analysis might be applied using guard cells as the tissue type.²³

The in vitro optical methods mentioned above may be implemented in vivo.

Uses of ordered water by the plant

The uses of callose by plants are essentially the uses of ordered water by plants. These uses will be illustrated by a set of hypothetical examples. The characteristic of ordered water given In Section C will be assumed valid and operational. Ordered water will be taken as strongly ice-like in its crystallinity.

Callose/ordered water within a pore: two levels of conductivity

Figure 3 shows a pore within which is an annulus of callose and an annulus of ordered water. This could be a sieve plate pore with a thickness of one micrometer The pore is not totally occluded. There is a space within the two annuli containing unordered water. The pore diameter is one micrometer. The callose layer is 0.05 micrometers in radial thickness and the ordered water is 0.2 micrometers in radial thickness. Assume that an electric field exist from one side of the pore to the other.

Figure 3.

Cross section and side view of the duality of a layer of callose and ordered water lining a sieve tube plate circular pore. The concavity limits the extent of the thickness of the ordered water Passage of ions other than protons and larger molecules is limited to the unordered water. Labels: CW, cell wall of circular pore; CA, callose; OW, ordered water. UW, unordered water. This diagram is not to scale.

The question is how fast and how many protons move through the pore. Alternately, does the ordered water enhance proton movement. What influence does the ordered water have on the movement of other ions.

The movement of ions through the pore is described by Eqn. 4.146 in Ref.10 . This equation relates ion velocity to the magnitude of the electric field. The mobility of the ion is the proportionality constant in this relation (See Tables 1 and 2 in Section C)

$$velocityoftheion=mobility\ast fieldstrength\left[meters/second\right]$$

$$v=\mu \ast F$$

Table 3 gives the results of applying this equation to the various particles passing through the one micrometer thick pore.

Table 3.

Transit time and velocity of particles passing through the sieve plate pore shown in Figure 3.

Particle	Transit time, Seconds	Velocity, meter/sec	Mobility, msquared/voltsec	Electric Field, volt/m
Proton in unordered water	3.3*10E−4	3*10E−3	3*10E−710	1*10E+4
Proton in ordered water	2*10E−6	5*10E−1	5*10E−510	1*10E+4
Potassium ion in unordered water	2*10E−3	5*10E−4	5*10E−810	1*10E+4
Sucrose	6*10E−3	1.6*E−414,18		Not applicable*

¹⁰ Table 5.2, ¹⁴ p170,¹⁸ p515, *Sucrose is not ionic.

The data in this table indicates that a proton moves through the ordered water 1000 times faster than a potassium ion moves through the unordered water. The proton also moves about 350 times faster than a sucrose molecule assuming no coupling between the proton and the sucrose. If coupling did exist, the proton movement would probably be slowed down and the sucrose movement would be sped up, see Section E5.

Wound callose: a blocking agent

Wound callose is ordered water in a region at a volume to preclude passage of ions through the region. The formation of callose in locations wherein it blocks passage may utilize ordered water only as a blocking agent for other ions and larger entities.

Callose/ordered water as a filter: plasmodesmata and pollen tubes

Under the callose/ordered water hypothesis, the entry point of the plasmodesmata into the lumen of the sieve tube is a volume or plug of ordered water such as shown in Figure 4.

Figure 4.

Schematic Diagram of a Plasmadesma containing callose/ordered water. The ordered water (in blue) rises above a layer of callose (not shown) in the xz plane and occupies the entire area of the passage. A desmotubule (not shown) passes through the plasmodesma in the xz plane. Bifurcations in the passage such as shown in Figure 6 begin in the z axis direction to the left of this callose/ordered water region. Only protons pass through this plasmodesma. This diagram is not to scale.

The ordered water is spread over the entire area in the xy plane. Only protons can pass through this block of ordered water. Ions other than protons are blocked. Larger entities are also blocked. This set of conditions holds only for plasmodesmata which contain sufficient callose and ordered water to totally encompass the opening in the manner of Figure 4.

Callose in Pollen Tubes are as example of a similar situation. The callose/ordered water deposition is described as a plug.⁵ But the plug under the callose/ordered water hypothesis is selective in that protons can pass through the plug but other ions cannot. The presence of high magnitude charge transfer into and out of the pollen tube has been documented by Weisenseel.²³ But the ion type carrying the charge was not reported. The putative presence of ordered water within the pollen tube would suggest that the charge carriers within the pollen tube measured by Weisenseel were protons.

Callose/ordered water in stomates: a protonic circuit

Callose (and putatively, ordered water) is present in the wall of stomatal guard cells.²⁴ Protons have been identified as the carrier of electrical current across the guard cell plasma membrane during patch clamp experiments. This suggests a protonic electrical circuit function of the ordered water. ^6,25 The circuit describing this situation is given in Figure 5. This is an example of a protonic circuit with a source of protons and an ordered water path to transfer these protons. The direction of proton movement shown is arbitrary and only illustrates the dynamics of the circuit. This example is relevant from several viewpoints. First, the callose is present as microfibrils. This supports the Author’s assumption of a linear configuration of the callose polymer discussed in Section B1 above. Second, the proton are present as a result of a proton pumping mechanism at the membrane.⁶ Proton emission into the apoplast is now coupled to proton transfer in the apoplast. Third, the protonic circuit is complete within a micro space scale. Protons are conserved. They have an origin and a destination. There is an equilibrium at the beginning and end of the proton movement. The circuit operates at a “micro” level.

Figure 5.

Stomatol Guard Cell and Callose/Ordered Water at its Surface Protons are pumped across the plasma membrane of the guard cell and into the region of the ordered water. They then move through the ordered water, back across the plasma membrane of the guard cell and into the smplast. The location of the dorsal and ventral wall and current direction are all arbitrary. Labels: O, Ordered Water; PM, Plasma Membrane; GC, Guard Cell; DW, Dorsal Wall; VW, Ventral Wall; PP1, Proton Pump1; PP2, Proton Pump2. Callose is shown by the purple strip. This diagram is not to scale.

The callose microfibrils are radial.²⁴ This would suggest that the ordered water, while isotropic, sustains radial proton movement within the isotropic ordered water.

The presence of a second proton pump is a requirement to move proton back to their point of origin. This assumes that there is a polarity reversal of an incremental electrical potential across the plasma membrane over the distance between the two proton pumps.

The guard cell and the callose layer on its periclinal surface comprise a protonic circuit one of whose components is a proton pump across the plasma membrane.

Proton microloop in the phloem: a supracellular protonic circuit

Existence of a proton microloop

Callose has been found to be a necessary part of phloem transport.²⁶ The definitive placement by the plant of callose in the pores of the sieve cell plates and along the walls of the plasmodesmata connecting the companion cells and the sieve cell lumen strongly suggests the plant has set up these locations as ordered water paths for high speed proton movement.¹⁷ The observation by Barratt and the locations of callose within the sieve tube/companion cell complex suggests that these locations are part of a proton microloop. Figure 6 illustrates the form of a proton microloop within the sieve tube/companion cell complex using callose/ordered water in two critical locations: in the lining of the sieve plate pores and in the entry region on the lumen side of the plasmodesmata. The form of the sieve plate pore and plasmodesmata are discussed in Sections E1 and E3 and shown in Figures 3 and 4.

Figure 6.

Schematic Diagram of a Proton Microloop in the Sieve Tube/Companion Cell Complex. The dashed black line indicates the path of protons around the microloop. The blue squares indicate the location of callose/ordered water. The green shading indicates a deficit in sucrose concentration at the end of Region 1–2 and an excess in sucrose concentration at the beginning of Region 3–4. Labels: CC2, Companion Cell2; CC3, CompanionCell3; PD2, Plasmodesmata2; PD3, Plasmodesmata3; CW, Sieve Tube Cell Wall; SPP, Sieve Plate Pore; ST, Lumen of the sieve tube. Region x-y is the length of lumen between PDx and PDy. The bifurcation in the plasmodesmata on the companion cell side is indicated by the expanded volume to the left of the ordered water. This diagram is not to scale.

Proton path defining the microloop

Consider the path of the proton movement. Companion cells 2 and 3 establish an electrical potential gradient in the lumen of the sieve tube between the cells. This is designated Region 2–3 in Figure 6. Protons move out of the symplast of Companion Cell2 through Plasmodesmata2 and into Region 2–3 of the lumen. The protons “attach” to sucrose molecules in the region and essentially drag the sucrose molecules through the lumen. When the proton/sucrose entity reaches Plasmodesmata3, the protons “detach” from the sucrose and move through the plasmodesmata and into the symplast of Companion Cell3. A buildup of protons occurs in the symplast of Companion Cell3. At this point in time, the electrical potential gradient ceases.

A second electrical potential gradient is now established between Companion Cell3 and Companion Cell2. Protons move across the plasma membrane of Companion Cell3 and into the apoplast between the companion cells. They cross the plasma membrane and return to the symplast of Companion Cell2. This completes the loop. Protons are conserved, that is, the concentration of protons at the beginning of the cycle is the same as the concentration at the end of the cycle. Energy for the proton movement is drawn from the companion cells. Sucrose has been transported along the lumen of the sieve tube.

Coupled conduction and diffusion

The movement of sucrose from the beginning of Region2-3 and buildup of sucrose at the end of region 2 causes an unbalance of sucrose concentration at both locations. This is shown in green in Figure 6. This sets up a diffusion of sucrose from Region 1–2 and a diffusion of sucrose into region 3–4. At the time and space scale of the conduction mechanism (milliseconds and micrometers) diffusion is quite rapid.^18,27,28 The result is a combined conduction/diffusion movement of sucrose extending over three regions of the lumen.

Significance of ordered water in this microloop

Ordered water is the sine qua non of this microloop. The callose/ordered water in the plasmodesma in the entry region of the passage between the lumen to the companion cell performs three necessary functions. First, consider that electrical potential gradients are not selective. All ions of both polarities move in response to the electric field. For example, potassium ions would be drawn out of the symplast of Companion Cell2. Even though their mobility is much less than protons, they do move. The ordered water in the plasmodesma blocks this movement. The same blockage holds for ions moving from the lumen of the sieve tube into the companion cell. This blockage prevents a serious unbalance of the ion concentrations in the companion cell symplast. Second, the ordered water will pass protons at a very enhanced level (See Table 3). Third, protons pass through in both directions in the plasmodesmata of the microloop. In other words, all three electrical characteristics of ordered water are utilized. Furthermore, if ions are blocked by the ordered water, larger entities such as pathogens will certainly be blocked.

Protons are attached to sucrose in the lumen. The microloop is concerned with these two molecules. Transit of other molecules in the lumen is ancillary to this movement. Gilbertson discusses such movement.²⁹

The movement of the sucrose is enhanced because it is now pulled though the pore by virtue of its attachment to the proton. This is in contrast to a purely hydraulic passage.³⁰

The method of attachment and detachment of the proton from the sucrose is not known. Nor is the stoichiometry of the proton/sucrose entity known. A coupling is only assumed. However, this theoretical example illustrates that protons, callose and ordered water can be a feasible part of normal sucrose movement within the lumen of the sieve tubes in addition to proton assisted loading and unloading of the sieve tubes themselves. The latter operation is well documented.^6,26,31

Callose/ordered water in mega and microsporogenesis

Temporal and spatial variation of the callose deposits

Callose has temporal variation, it begins in the premeiotic and disappears in the post tetrad phases.³² Callose has spatial variation, it varies from the chalazal pole to the micropylar pole.³³ Under the duality hypothesis, this indicates the ordered water and its electrical properties also have a temporal and spatial variation.

Holes in callose walls

There is a distinct progression of changes in the wall between sporocytes during microsporogenesis.^32,34,35 Plasmodesmatal connections disappear and are replaced by a massive deposit of callose between the sporocytes. The callose wall contain holes ranging from 0.5 to 1.5 micrometers in diameter. They are not as symmetrical as a round pore.³⁶ But the perimeter is callose. As such they are similar in size to the pore shown in Figure 3. The result is a sequence of channels from one microsporocyte to the next. The electrical resistance between two microsporocytes 400 micrometers apart was measured at a level of 1.8* 10E+4 ohms.³⁷ This resistance was considered “low.” But the criterion that led to the term “low”was not reported. Esau concludes that these holes permit rapid transfer of materials through the sequence of sporocyte callose walls.³⁴ Under the duality hypothesis, the rapid transfer would be facilitated by an annulus of ordered water in each channel. The presence of ordered water per se does not insure rapid movement of materials through the channels. In addition, a driving force must be present. The situation is similar to the movement of sucrose through the callose lined sieve plate pores shown in Figure 3. But the driving force in this case is not defined.

Reason for the presence of the callose wall

According to Heslop-Harrison, “The most reasonable interpretation of the function of the callose wall is that it acts as a molecular filter, permitting the passage into the spores of basal nutrients but excluding larger molecules”.³³ The duality hypothesis would suggest that the filtering action comes from the layer of ordered water adjacent to the callose. The complex variations of callose and ordered water within the microsporocyte and megasporocyte over time and space would suggest that the proton property of the ordered water might also come into play. In other words, one might be observing changes in metabolic function in the form of proton microloops in the region of each individual spore during development.

Summary of the uses of callose/ordered water

These examples serve to illustrate possible uses of callose/ordered water in plant activity. The callose/ordered water is used at times for its physical properties and at other times for its electrical properties. Sometimes the plant elects to use plasmodesmata containing ordered water and at other times the plant elects to use a pore with varying areas of ordered water.

The examples also serve to point out that the presence of ordered water and the enhanced proton movement that is possible only gives rise to the requirement that a complete protonic circuit must be defined including a driving force.

From a broader perspective, the duality of callose and ordered water utilizes two commonly available materials, that is, glucose and water, to achieve crystallinity at normal environmental temperatures. This is a remarkable synthesis.

Overview: protonic circuits and electronic circuits

Movement of protons through ordered water in plants is an example of a protonic circuit. Algae and land plants made use of what was readily available beginning with the so-called green lineage over 600 million years ago.⁴ This is in contrast to Man’s use of movement of electrons through refined materials such as copper and semiconductors beginning about 200 years ago.

“…and the seed sprouts and grows, and he knows not how it happens“ Mark 4/27

Funding Statement

This work was supported by the Agricultural Electronics Corporation.

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.