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Progress in Biophysics and Molecular Biology 89 (2005) 18



Hypothesis: the origin of life in a hydrogel environment

Jack T. Trevorsa,, Gerald H. Pollackb,1

Laboratory of Microbial Technology, Department of Environmental Biology, Ontario Agricultural College, University of Guelph, Guelph, Ontario, Canada N1G 2W1 b Department of Bioengineering, Box 357962, University of Washington, Seattle, WA 98195, USA Available online 22 September 2004

Abstract A hypothesis is proposed that the rst cell(s) on the Earth assembled in a hydrogel environment. Gel environments are capable of retaining water, oily hydrocarbons, solutes, and gas bubbles, and are capable of carrying out many functions, even in the absence of a membrane. Thus, the gel-like environment may have conferred distinct advantages for the assembly of the rst cell(s). r 2004 Elsevier Ltd. All rights reserved.
Keywords: Assembly; Bacteria; Cell; Clays; Cytoplasm; Division; Earth; Evolution; Growth; Hydrogel; Life; Membrane; Origin; Water

Contents 1. 2. 3. 4. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 The hydrogel environment: superior to a liquid environment for the origin of life? . . . . . . . . . . . . . . . . . . . . 2 Gels and work production . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 Metabolism in gels. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4

Corresponding author. Tel.: +1-519-824-4120 ext. 53367; fax: +1-519-837-0442.


E-mail addresses: jtrevors@uoguelph.ca (J.T. Trevors), ghp@u.washington.edu (G.H. Pollack). Also to be contacted for correspondence.

0079-6107/$ - see front matter r 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.pbiomolbio.2004.07.003

2 5. 6. J.T. Trevors, G.H. Pollack / Progress in Biophysics and Molecular Biology 89 (2005) 18 Transition from pre-cell to cell . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 Conclusions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

1. Introduction The origin and evolution of life have been long debated, with fundamental questions still unresolved. What, for example, were ancient lifes most common single-celled forms (Schopf, 1999)? For what biological, chemical, or physical life signatures ought one to be searching (Bebie and Schoonen, 1999; Bashkin, 2002; Cairns-Smith, 1985, 2001; Deamer, 1997, 1992; Des Marais and Walter, 1999; Gold, 1992, 2001; Ingber, 2000; Morchio and Traverso, 1999; Morita, 1999; Nilson, 2002; Segre et al., 2001; Trevors, 2001a, b, 2002a, b, 2003ac; Trevors and Abel, 2004; Trevors and Psenner 2001; Wachtershauser, 1997)? Where should one look for ancient life remnantsin thermal springs, deep ocean vents, subsurface cores, deep hot oily subsurfaces (Gold, 1992, 2001) where water and oil form interfaces, or in available extraterrestrial samples? In searching for answers to these questions, an implicit presumption is often made that the cell is an aqueous suspension of solutes. The pre-cell, then, is also considered to be an aqueous suspension, eventually surrounded by a membrane to become a cell. However, it is known that the cell is a gel (Pollack, 2001). With the cytoplasm treated as a gel rather than as an aqueous suspension, a question worth considering is whether the various conundrums that have limited progress in understanding the origin of life and the genetic instruction set might be better resolved. We propose that a primitive hydrogel was a more suitable environment for the assembly of precells, and then cells capable of growth and division. Gels, for example, retain their integrity even in the absence of a membrane. Primitive organelles, nutrients, ions, proteins and nucleic acids could remain ordered and in continuous, close molecular physical proximity within the gel without the danger of dissipating, as it would in a strictly aqueous environment with free diffusion. Hence, the question of how the pre-cell with no membrane and a very small mass could retain its integrity, need not be an issue with the cytoplasm viewed as a cohesive gel. Further, many basic functions performed by the cell or pre-cell are capable of being carried out by gels themselves, as we elaborate below.

2. The hydrogel environment: superior to a liquid environment for the origin of life? Although gels are employed for numerous functions from absorbing urine in disposable diapers, and for immobilizing enzymes and cells, to controlling drug release in time-release pharmaceuticals, there is no agreed-upon denition of a gel. A textbook denition is that the elastic modulus is greater than the viscous modulus: unlike liquids, gels do not ow as a result of steady shear. A phenomenological denition is that gels are solid or solid-like materials that

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consist of two or more components, one of which is a liquid of some abundance (Almdal et al., 1993). One feature of hydrogels, i.e., gels in water solvent, is that the solvent is not ordinarily bulk water. As dipoles, water molecules will adsorb onto hydrophilic surfaces, and onto one another, forming networks (Ling, 1965). When enmeshed in a polymer network, such layered, or structured water holds together the network, forming a stable gel (Pollack, 2001). Effectively, the hydrogen bonding in water can hold together even polymers that are not cross-linked to one another, as in many common colloidal gels such as gelatin (Bella et al., 1994; Bromberg and Ron, 1998). One feature of this kind of structured water is that solutes are excluded. Exclusion is largely size based. Because of both enthalpic and entropic considerations, the larger the solute the more profoundly the solute will tend to avoid the structured water environment inside the gel/cell (Negendank, 1982; Ling, 1984; Wiggins, 1990). Similar size-based exclusion of solutes occurs immediately outside cells/gels, for distances in the order of 100 mm or more (Zheng and Pollack, 2003). For the pre-cell or primitive cell and even the modern cell, the solute-exclusion principle has important consequence. It provides a mechanism for partitioning of critical ions between the inside and outside. Thus, sodium, with a large hydrated diameter of 5 A, will be kept from entering the cell/gel in any quantity, whereas potassium, with a smaller hydrated diameter (3.8 A) and high afnity for protein surfaces (Edelmann, 1988; Cameron et al., 1996; Kellermayer et al., 1994), will enter more easily and accumulate. The principle is commonly exploited in gels known as ion-exchange resins. This property may have provided a mechanism by which the pre-cell, without a membrane, established the required ion-concentration gradients. Channels and pumps are not needed; the cytoplasmic gel matrix itself is sufcient. Indeed, energy is not needed; because this is an equilibrium process, the requisite separation is maintained indenitely, without a continuous supply of energy. A continuous energy supply may or may not have been present when the rst pre-cells were being assembled. If it was, it remains unknown if this energy was readily capturable until mechanisms for energy capture, production, usage, and storage became available. Hence, ion partitioning as an equilibrium process had distinct advantages.

3. Gels and work production Unlike the common perception of being largely inert, gels are capable of doing work. The central mechanism of work production is the polymer gel phase transition. Postulated by Dusek and Patterson (1968), and amply conrmed by the late Toyo Tanaka (1981) and subsequently by many others, the polymer gel phase transition is a critical phenomenon much like the waterice transition, in which a small change of environment results in a major, discontinuous, change of physical structure. Volume changes of 1000 times are not uncommon (Osada and Gong, 1993); one state is highly hydrated, the other not. These changes of volume (or length) are capable of performing work, as for example in articial muscles (Bar-Cohen, 2001). Such work could have been used by the pre-cell for metabolism and other critical processes, as it is used in diverse intracellular organelles today (Verdugo et al., 1992; Tasaki, 1999; Pollack, 2001).

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Energy for any such transitions would have been derived from the sun, either directly or indirectly. Sun-induced evaporation, for example, could concentrate solutes, shifting the equilibrium towards polymerization and thereby creating conditions more favorable for polymer growth. Triggers of the phase transition include any of a vast number of environmental shifts, ranging from ion content, to pH and temperature. Thus, Sun-induced heating could raise the temperature sufciently to trigger a transition, which could then perform work. Light itself can sometimes trigger phase transitions (Hoffman, 1991). Hence, work-producing phase transitions would have been driven by the main source of energy, the sun. Phase transitions have been demonstrated to produce various kinds of work relevant to cellular function. While the earliest pre-cells may have relied on diffusion alone for nutrient ow, the early presence of a simple mechanism for convective ow would have enhanced the versatility of the pre-cell or early cell. Convective ow can be produced by simple propagating phase transitions; this occurs along actin laments, microtubules, and presumably also along similar polymers of a more primitive nature that may have been involved in early cell division (Pollack, 2001). Moreover, polymer-gel phase transitions are centrally implicated in basic cellular processes such as secretion, potential changes similar to action potentials, and various types of movement. All of these functions can be accomplished by biopolymer gels, and frequently also by articial gels that mimic what happens biologically (Pollack, 2001). Thus, polymer-gel phase transitions offered a simple mechanism by which the pre-cell or early cell(s) could have accomplished various simple tasks.

4. Metabolism in gels The hydrogel environment provided a matrix conducive to non-enzymatic chemical and enzymatic reactions. Enzymatic reactions can occur in narrow, nanometer-scale pores, where water is almost certainly structured (Wiggins, 1990). Thus, a gel environment may have provided a more stable environment, while at the same time allowing chemical and eventually enzymatic reactions to occur, with enzymes remaining active for longer periods of time. Any enzymes necessary for assembly of the rst cell(s) that remained active longer would have been more useful than enzymes with a rapid turnover, especially if the enzymes were synthesized slowly and/or were slow acting with the ability to catalyze the transformation of numerous substrates (Demetrius, 1988). Central to the assembly of the rst cell(s) would have been the mechanisms of energy production, storage and utilization. What advantages would a gel-like cytoplasm have offered for these processes, compared to an aqueous cytoplasm? First, in the pre-cell(s), all components would be held in a exible but not too viscous gel that physically maintained components in contact with one and other over long periods of time. Without physical contact, reactions cannot proceed. Further, the gel state brought some order to an otherwise disordered environment, was simpler to encapsulate with an evolving cytoplasmic membrane, and also concentrated solutes necessary for the evolution of cellular metabolism. The gel environment also facilitates processes such as growth and division. A cohesive gel-like cytoplasm is an easier environment to partition into two entities without cytoplasm streaming away. The mechanism that divides a single bacterial cell into two offspring cells, for example,

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accomplishes this amazing activity with no loss of genetic material or cytoplasmic contents. The hydrogels cohesive nature would keep cellular components together. In addition, the gel nature of cells allows diffusion of solutes and gases (e.g., eventually oxygen for aerobic respiration) to move in and out without expenditure of energy. This type of diffusion is also possible in an aqueous environment; however, once the gases entered the gel environment, their diffusion out of the gel will have been slowed, increasing their availability for evolving metabolism. Gels may have contained various gases useful for metabolic reactions, such as N2, H2, CH4 and CO2, which were likely ubiquitous on the primitive Earth. Hydrogen encapsulated in a gel matrix may have been an early substrate for primitive enzymes. Present day bacteria utilize hydrogen in some reactions, and coliform bacteria can utilize hydrogen as an energy source but not for growth; growth requirements are now fullled by other substrates (Morita, 1999). In summary, the gel environment could have trapped and concentrated gases as they entered the gel, retarding their loss and concentrating them, while at the same time making them available as substrates in the rst cell(s). A hydrogel cytoplasm may also have been conducive to the assembly of the rst macromolecules. The rst gel cytoplasm would have been much simpler as only some dissolved gases, ions, molecules, small peptides, proteins and catalytic RNA were likely to have been present. Proteins and nucleic acids would still, however, be capable of folding and taking on their structural conformations in the non-restrictive gel, whereas a strict aqueous solution would have offered less stablity, leading to decomposition. The ability of DNA to reside as linear DNA or closed circular DNA in present day agarose gels illustrates that a gel environment used in electrophoresis still allows the DNA to take on different congurations, the DNA remaining functional for cloning experiments and polymerase chain reaction (PCR) analysis. DNAs stability and functionality are not affected in an adverse manner by the hydrogel environment. It is even possible that the DNA remains more stable. Gels are also used as a non-restrictive matrix for protein and RNA analysis without evident damage to these macromolecules. The gel matrix may have provided a stable environment for hundreds of millions of years that was not possible in a simple aqueous medium, where stability was challenged by turbulence, shear forces, and rapidly changing temperature, pH, and chemical concentrations, as well as by hydrolysis reactions. Unless environmental conditions lie within restricted ranges of pH, ionic concentrations and temperature, nucleic acids and mRNA are unstable during extraction from cells (Trevors and van Elsas, 1995). Even then, degradation can occur in seconds to minutes. This liability implies that the rst catalytic RNA must have been stabilized in a protective environment. Moreover, even a simple pentose such as ribose is difcult to make, and also unstable, as are ribonucleosides. Cytosine is very difcult to synthesize even in laboratory conditions (Shapiro, 1999). By maintaining a suitable pH, osmotic balance and temperature, a stable hydrogel cytoplasm may have minimized this difculty.

5. Transition from pre-cell to cell The transition between a primitive and a more complex pre-cell may have been the time the cell membrane was rst assembled. At some point through an unknown mechanism, the cell surrounded itself by a lipid bilayer, thereby dening the cellular entity. The advent of the cell

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membrane brought a conundrum. The appearance of the continuous cell membrane presented a largely impermeable barrier between the inside and outside. How then could nutrients pass from outside to inside, and waste products in reverse? Accommodating trans-membrane solute ow must have required the presence of membranous components similar to, but evidently more primitive, than current membrane pumps and channels. The problem is this: with no evolutionary pressure beforehand, by what mechanism could the machinery have arisen for the construction of these processes? The pumps and channels were needed instantly, to avert cell death. But development of the manufacturing capability would have taken eons. How could the membraneenveloped cell have communicated with the environment in the interim? The gel construct provides a basis to escape from the horns of this dilemma. The central point is that the gel holds itself intact whether a continuous membrane is present or not; it tolerates even large supercial orices. Such orices, by contrast, are not easily tolerated by a cytoplasm that is an aqueous suspension, as solutes would ow down their respective concentration gradients out of the cell. The cell would quickly perish. Even todays cells are leaky. The membrane contains a high density of proteins, as much as 80% by weight in bacterial cells and 50% in mammalian cells. Thus, the membrane is by no means a pure lipid bilayer. It is riddled with proteins that provide ample leakage pathways. Evidence for this is found in the fact that when additional leakage pathways are created (by even gross membrane violations), routine activity can often continue even though the membrane does not appear to re-grow (Maniotis and Schliwa, 1991; Yawo and Kuno, 1985; Albrecht-Buehler, 1980). One example is that of cells cut into two fractions by the tip of a micropipette. Despite this violation, the cell does not perish (Maniotis and Schliwa, 1991). In fact, the fraction with the required organelles goes on to divide and produce offspring. This example is but one of a halfdozen classes of decapitated cells that nevertheless continue to function (Pollack, 2001). Such seemingly anomalous behavior is anticipated within the gel construct. For vital processes to proceed, membrane continuity is unnecessary. Whether continuity is breached by the presence of ordinary leaky proteins, or by manual insult, cell functions need not be seriously affected. The gel holds together, allowing processes to continue. Hence, within the gel paradigm, the abrupt appearance of the insulating lipid-bilayer membrane presents no conundrum if the bilayer contains orices such as proteins to facilitate communication. There is no instant requirement for pumps, channels, or active transport mechanisms. Leaky orices may present challenges for aqueous suspension models, but not for gel structures.

6. Conclusions A hydrogel gel environment provided several advantages for the origin of life. It provided a stable environment not only for the accretion of polymeric mass, but also for subsequent cell division and evolution. By undergoing phase transition, hydrogels have the capacity to carry out various energy-requiring tasks, and there is evidence that phase transitions are used routinely today in highly evolved eukaryotic cells (Verdugo et al., 1992; Tasaki, 1999; Pollack, 2001). Furthermore, the gel paradigm offers a solution to the long-held conundrum of how the pre-cell coped before the abrupt appearance of the continuous cell membrane. We suggest that the

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recognition of the hydrogel-like nature of the cell needs to be further explored, as it may provide a fuller understanding of how the rst cells assembled and divided.

Acknowledgements JTT was supported was by a NSERC (Canada) Discovery Grant. GHP was supported by NIH Grant # 1 R01 AR44813.

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