A Chemical Composition to Make Them Cower: A Dive Into the Structure and Protective Adaptations of Sea Sponges
Olivia Ghiaur, Camille Heaney, Chris Hu, Vadim Zamaruyev
Abstract
Sponges have several feeding modes and use a variety of food sources, which underlies their key role in the cycling of several elements and importance in the foodweb. Sponges also produce a variety of toxins that they use in allelopathy against competitors, and in camouflage and defense against predators. The biomineralization of sponge skeletons has an interesting evolutionary history, is a multi-step process, and confers sponges a remarkable morphological plasticity. As a result, these animals can change their shape to adapt to their environment as they grow. Apart from adaptations of biological importance, the porous structure of spongin is a source of inspiration for new filtering and catalytic biomimetic devices.
Introduction
With so many evolutionary niches to fill, Porifera utilized the power of chemistry, allowing them to not only change their environment but change themselves (Valery et al., 2003). Each new interaction opened the door for a new adaptation. The abundance of silicon and oxygen led to the evolution of silicatein (Fig. 19) and collagen (Fig. 23). The latter two, along with spongin, drove the emergence of different spicules incorporated in bodies of increasing size and complexity through the cooperation of various specialized cells. The resulting phenotypic plasticity opened new avenues of interaction between sponges and other species. Sponges became prey and predators. These interspecific interactions triggered further chemical diversification. Sponges developed molecules for camouflage and defense and became important middlemen in the flow of chemical elements. In other words, chemistry sparked diversity (Valery et al., 2003).
Consumption
Sea sponges have adapted to living in environments ranging from nutrient poor to rich, light to dark, and with or without symbionts. As such, sponges have developed many pathways by which to take in nutrients from their surroundings (Valery et al., 2003).
Diffusion
Like all cells, the external, lining cells of sea sponges are capable of diffusion from the water directly into the cell. Diffusion occurs when there is a high concentration of a dissolved material in one medium (either gas or liquid) and this medium is separated from another by a boundary (in the case of sea sponges, a cell membrane). The dissolved material filters through the boundary until the external and internal mediums have the same concentration of material (Bailey, 2019).
Pinocytosis
Pinocytosis (Fig. 1) refers to the formation of a pocket to engulf extracellular fluid. As opposed to diffusion, this process is more controlled, occurring with larger or less polar molecules which cannot enter channels in the phospholipid bilayer. Instead, the cell bilayer folds in on itself, holding the polar medium and its content with the polar head of the bilayer. This pocket pinches off inside the cell creating a vesicle. From here lysosomes are deployed within the vesicle to break apart the nutrients into sea-sponge friendly packages. The remainder is handled by exosomes and released from the cell (Britannica, 2014).
Fig. 1. Pinocytosis [Adapted from Karpińska et al., 2022]
Phagocytosis
Phagocytosis enables unicellular organisms to consume particles larger than 0.5 µm in diameter (Fig. 2). Particles are sent through the ostia of sea sponges into choanocyte chambers. Choanocytes have flagella which allow them to “hold” microorganisms in one place (Steinmetz, 2019). Upon sensing the particle, choanocytes signal the remodel of the cytoskeleton and phospholipid bilayer to extend to cover the particle. Once these membranes close, a phagosome is formed. During maturation, this distinct vesicle binds with lysosomes and other hydrolytic enzymes to produce reactive oxygen species (like superoxide), and becomes very acidic (pH around 4.5). These work together to degrade the ingested material (Uribe-Querol & Rosales, 2020).
Fig. 2. Stages of the mechanism of phagocytosis. Blue arrows indicate the direction of the steps. (Ojas A. Deshpanda, 2023)
Carnivory
In the class Demospongiae, the family of Cladorhizidae adapted to fit environments with far less nutritional content. Appearing as a sea sponge “skeleton”, this class lacks the classic poriferan aquiferous system but maintains the siliceous spicule structure. The spicules have evolved hook-like qualities, used to entangle the legs of small crustaceans.
Digestion is done by migrating elongate cells, which specialize from stem cells at the base of the sponge structure. A few hours after the prey have been captured, a layer of pinacocytes form around the prey. Then archeocytes and bacteriocytes concentrate around the prey creating a cyst-like structure on the sponge. Within a week, identifiable nutrients from the prey are phagocytosed and the undigested elements (like carapaces or gills) are released (Vacelet & Duport, 2004).
Symbiosis
Porifera do not function isolated from other organisms, they have symbionts that are paired with them through a plethora of systems, including but not limited to the dissolution of carbon and nitrogen, vitamin biosynthesis and toxin formation (Carrier et al., 2022).
In clear shallow waters, sea sponges work with photosynthetic symbionts, like zooxanthellae and cyanobacteria. These use the energy of photons to excite electrons forcing them to create glucose and ATP from water and carbon dioxide (AKA photosynthesis). This is a vital pairing because it directly contributes to the ability of sea sponges to fixate dissolved carbon, by receiving it in the form of glucose from the microbial symbionts.
Deep-sea sponges pair with chemoautotrophic bacteria to help fix Carbon dioxide (CO2). Specific ammonia-oxidizing archaea, like Nitrosopumilaceae, fixate carbon in the form of the 3-hydroxypropionate/4-hydroxybutyrate cycle (Garritano et al., 2023). The 3-hydroxypropionate/4-hydroxybutyrate carbon fixation cycle (Fig. 3) takes dissolved CO2 from the water and synthesizes it into acetyl CoA. This process is more efficient than sponge diffusion which is critical in nutrient deficient waters (Loder et al., 2016).
Fig. 3. 3-hydroxypropionate/4-hydroxybutyrate carbon fixation cycle (Loder et al., 2016)
There are two overarching groups of sponges in relation to symbionts: low microbial abundance (LMA) and high microbial abundance (HMA). These are important distinctions because sea sponges are a collection of cells rather than tissue-based, meaning that the symbiont microbes are incorporated within the cellular structure of the host. This relationship is so integral that about 50% of sponge biomass is made of their bacterial symbionts (Kiran et al., 2018).
Nutrition
In the carbon, nitrogen, and phosphorous, cycles sea sponges function as sources, sinks, and cyclers (Valery et al., 2003).
Oxygen
While H2O is polar and O2 is nonpolar, dissolved oxygen (DO) is critical to almost all aquatic life. Oxygen is able to dissolve in water due to the hydrophobic effect, but in the oceans, oxygen mainly comes from the release of photosynthetic organisms (Mader et al., 2017).
Sea sponges use oxygen for respiration to create ATP, which powers cellular activity.
Carbon
There are two overarching types of carbon which sea sponges ingest, dissolved organic matter (DOM) and particulate organic matter (POM).
DOM is largely inaccessible to microorganisms, but exists in bodies of water in the form of glucose, amino acids, etc. Sponges are uniquely capable of taking up this matter directly via pinocytosis (Rix et al., 2020) and receive the synthesized form of DOM from their carbon fixing symbionts.
Sponge ingested POM can be alive, in the form of pico- or nano-planktonic cells, or non-living, in the form of detritus. For sponges this digestion process occurs via phagocytosis.
The use of carbon in sea sponges can be summarized as (I)ngestion, biomass (P)roduction, (R)espiration, and (E)xcrement, which relate as follows:
Equation 1.
I = P + R + E
Depending on temperature, pumping activity, and physiological processes, such as growth and reproduction, the respiration rate of sponges can be between 0.21 to 24.6µmol O2cm-3h-1 (Maldonado et al., 2012). These varied respiration rates are related to the metabolic efficiency of the sea sponge by the ratio between ingestion and respiration. Thus, at times when there is high carbon availability and they are internally active, sponges are at their most successful.
The extra carbon is released from the sponge system in the form of detritus. This is crucial for the benthic ecosystem (Fig. 4) as it allows other macro-organisms to use the fixated carbon, which they cannot access in the open water, and further contribute to the cycle.
Fig. 4. Carbon cycling surrounding sea sponges. POCdet represents detrital POC (Maldonado et al., 2012).
Nitrogen and Phosphorous
Two other nutrients that are vital in metabolic processes are nitrogen and phosphorus.
Marine sponges are capable of taking in dissolved inorganic nitrogen and phosphorus, like NH4+ , NO3–, and PO43- , through diffusion and pinocytosis, and organic forms of nitrogen, like amino acids, primarily through phagocytosis. However, this process is quite inefficient, and symbionts are much more competent in this task. Symbiotic bacteria, archaea, and fungi have all been regularly found associated with sponges as ammonia-oxidizers, nitrite-oxidizers, and denitrifiers (Fig. 5.) (Karpińska et al., 2022).
Fig. 5. Nitrogen fixation in marine sponges (Kiran et al., 2018).
This association is integral for sea sponges as nitrogen and phosphorus are necessary for protein and nucleic acid production, but it is also necessary for the surrounding flora and fauna. The importance of porifera in the nitrogen cycle has been found in sea habitats from temperate to tropical. This is because they act as a source of dissolved organic nitrogen (Fig. 6) for pelagic environments and are players in the remineralization of organic nitrogen (Bell, 2008).
Fig. 6. Nitrogen and phosphorous cycle at the poriferan level (Maldonado et al., 2012)
Effect on ecosystem
The role of sea sponges as keystone species in their habitats is common because they do not just physically reshape an environment, they nutritionally reshape it as well.
Benthic (those on the base of the sea) and pelagic (open ocean) environments are connected. Their coupling contributes to various biogeochemical cycles, as benthic fauna are important in organic matter retention and remineralization, while pelagic organisms produce fresh organic matter (Ehrnsten et al., 2019). Sea sponges participate in this relationship as their pumping system sends material upwards, facilitating benthic-pelagic exchange and bioturbation on the sea floor (Bell, 2008). They also function as an intermediate between dissolved and microbial nutrition, and higher levels of the food chain.
Chemical defenses of sponges
Sponges are sessile, which means they are rooted in the same location their entire lives. Sponges cannot escape predators. As a result, they have evolved to have chemical defense mechanisms. Sponges contain an abundance of secondary metabolites that protect them from predators, prevent fouling of other organisms, and compete for space in crowded environments. As sponges filter water, they can digest microorganisms or retain them within their porous bodies. Some of these microorganisms are involved in the biosynthesis of the natural products of sponges which enables chemical defenses. Sponges prevent fouling by having clear pathways for water filtration, they can conquer densely populated areas with their metabolites, and they have mechanisms to produce potent toxins without destroying themselves (Varijakzhan et al., 2021).
Additionally, the environment of the sponge has an influence on the accumulation of toxic constituents within the sponge. Sponges that live in tropical environments have more chemical compounds made for defense than sponges that live in colder environments. Tropical environments have more intense competition and feeding pressures which has resulted in tropical sponges containing more toxins (Proksch, 1994).
Allelopathic ability
Allelopathy is when toxins are secreted to prevent the growth of other organisms around the sponge, also allowing sponges to secure space for their larvae (Singh & Thakur, 2016). Sponges grow in environments that are crowded with other sessile organisms. They have developed the chemical defense mechanism of allelopathy to secure their spots in the limited benthic habitat. The allelochemical properties of sponges and other organisms are not well understood. The interactions can be through direct contact or the release of chemicals into the environment around the sponge (Ternon et al., 2016).
One example of an allelopathic sponge with a direct toxic effect is Cliona tenuis (Fig. 7). This sponge produces clionapyrrolidine A [(−)-(5S)-2-imino-1-methylpyrrolidine-5-carboxylic acid], a compound that, upon contact, kills coral tissue and reduces the photosynthetic efficiency of certain corals. The toxin only works through forced contact, this implies that the contact that would result in coral death can only happen during a storm. (Chaves-Fonnegra et al., 2008).
Fig. 7 Image of Cliona tenuis from Grand Cayman (Charpin, 2023).
Another example of an allelopathic sponge is the Crambe crambe which releases chemicals into the seawater. C. crambe synthesizes the bioactive polycyclic guanidine alkaloids (PGA). There are two families of PGAs: the first family, crambescins, contains two cyclic rings, and the second family, crambescidins, contains five cyclic rings (Fig. 8). The sponge contains high concentrations of these compounds, which may be involved in the chemical mediation of other marine micro and macro-organisms. Additionally, the water surrounding the sponge contained PGAs, which means that the sponge has a continuous transfer of metabolites into the seawater. The sponges can release chemicals into the environment by creating spherulous cells that contain spherules filled with metabolites. The spherulous cells exit the sponge through the oscula and burst creating a shield of metabolites around the sponge (Fig. 9). It is demonstrated that PGA compounds cause defects in the embryos of sea squirts. Sea squirts are other sessile creatures competing for space, and C. Crambe prevents their development, proving its allelopathic abilities (Ternon et al., 2016).
Fig. 8. The Chemical structures of two ring Crambescins A, Crambescins B, and Crambescins C. The chemical structure of five ring Crambescidins (Ternon et al., 2016).
Fig. 9. Schematic of how the chemical shield is formed by C. crambe. Left green circle shows crambescins and right red circle shows crambescidins (Ternon et al., 2016).
Antifouling ability
Sponges settled on the sea floor do not want other organisms such as algae, barnacles, ascidians, and bryozoans to settle on top of them and evolved anti-fouling abilities. Biofouling is a multistep process that allows larger organisms to stick to underwater surfaces (Fig. 10). The process begins with a conditioning film where an organic particulate layer is formed by proteins, carbohydrates, and glycoproteins. The surface is then conditioned for microorganisms to stick through electrostatic and Van der Waals forces. Then, microorganisms will stick to the surface as they naturally pass by the surface via the movement of water. Next, the microorganisms will secrete an extracellular polymeric substance anchoring them to the surface. This creates a gel matrix that allows for enzymatic interactions, nutrient exchange, and environmental stressor protection. The initial soft macro-fouling allows for the growth of hard macro-fouling of organisms; starting with small eukaryotes and followed by large eukaryotes. (Müller et al., 2013). One metabolite used in sponges to prevent the settlement of the barnacle Balanus improvisus is agelasine D (Fig. 11) (Sjögren et al., 2008). An additional compound is ageloxime D. Agelasine D has antimicrobial properties and Ageloxime D inhibits the biofilm of the barnacle. (Hertiani et al., 2010).
Fig. 10 The basic biofouling process stages (Müller et al., 2013). Stages 1-3 create the biofilm for larger organisms to later attach.
Fig. 11. Chemical structure of Agelasine D. [adapted from (from Sjögren et al., 2008)]
Symbiotic relationship to produce toxins
As filter feeders, sponges take in a lot of water as well as the microbes in the water. Some sponges have symbiotic relationships with the bacteria that enter their bodies. In particular, the sponges of the order Dysideidae have a cyanobacteria living within them that produces polybrominated diphenyl ethers (PBDEs). The natural PBDEs are remarkably like the toxic human-made brominated flame retardants (Fig. 12a & b)(Agarwal et al., 2017). The PBDEs of the sponges are used as a chemical defense against predators. The mechanism by which they are toxic against other organisms is unknown. However, they provide the sponge with some microbial resistance. The PBDEs created by the cyanobacteria tend to accumulate and crystallize at high concentrations in the ectosome tissues of the sponge. This could be how sponges avoid the toxic effects of the compounds(Poston & Saha, 2019).
Fig. 12. The similarity in the chemical structure of polyhalogenated molecules as synthetic toxins (a) and naturally produced marine toxins (b) (Agarwal et al., 2017).
Another example is from the sponge Discodermia calyx (Fig. 13a). The Sponge has a symbiotic relationship with the bacteria Candidatus Entotheonella. The bacterium within the sponge produces a toxin called Calyculin A. The structure of the toxin contains tetraene, oxazole, phosphate and dimethylamino groups. The toxin is synthesized via a hybrid pathway of polyketide synthase (PKS) and non-ribosomal peptide synthase (NRPS). Then, there are modifications such as nitrile formation to finish the structure. This toxin works by inhibiting the protein phosphatase 1 and 2A (Wakimoto et al., 2014). These proteins are important in the regulation of cell functions such as cell signaling, metabolism, cell cycle transitions, embryonic development and stress response (Courtney & Deiters, 2019). The disruption in these functions from the toxin can lead to cellular death. When the sponges work together with the bacteria, they can have a chemical defense from Calyculin A (Wakimoto et al., 2014).
Fig. 13. (a) The sponge Discodermia calyx. (b) Structure of the toxin, calyculin A (Wakimoto et al., 2014).
Discodermia calyx can use this toxic compound to defend itself. However, the toxin works by targeting enzymes that are important to all eukaryotic cells. To combat this, the relationship with the bacteria has evolved to have a mechanism to prevent self-toxicity. The mechanism used by the sponge and the symbiotic bacteria is to store the toxin as a derivative, phosphocalyculin A. Phosphocalyculin A has an additional phosphate group this diminishes the toxicity since the single phosphate group on Calyculin A is responsible for the inhibitions of protein phosphatases 1 and 2A. When the sponge tissue is disturbed, the bacteria will release phasphatase CalL which will activate the toxin by converting phosphocalyculin A into Calyculin A (Fig. 14). This mechanism allows the sponge to contain the toxin within itself without being harmed and rapidly activate the toxin when needed (Jomori et al., 2021).
Fig. 14. Phosphate-activated chemical defense mechanism of conversion of Phosphocalyculin A to Calyculin A (Jomori et al., 2021).
Silica Biomineralization in Sponges
The earliest fossil evidence of sponges dates to the Ediacaran (542–580 Ma) the latest period of the Proterozoic era preceding the Cambrian (McMenamin, 2005). In early Cambrian ecosystems, sponges were one of the most diversified groups. They were the second most diverse group in the Chengjiang biota (520-530 Ma) where both demosponge and hexactinellids were identified (Fig 15). A key innovation underpinned the ecological success of these animals: their skeleton. A skeleton provides a species with the opportunity to scale up and develop a complex organization. The skeleton of sponges relied on two key molecules: collagen which formed the extracellular matrix and silicatein which produced spicules. Collagen needed oxygen, which was abundant since the great oxidation event that happened 2.4 billion years ago (Olejarz et al., 2021). Silicatein needed silicon (Muller et al., 2011).
Fig. 15. Evolution of sponges. The earliest sponge fossils are found in the Ediacaran biota. Fossil evidence from Chengjiang and Burgess Shales supports their diversification during the early Cambrian period. Sponges evolved in a silicon-rich ocean. Notice that more recent events are to the left. (Muller et al., 2011).
Silicon is the second most abundant element in Earth’s crust, and its concentration was significantly higher in the Ediacaran oceans than today (Tyler, 2013). These concentrations were attributable to two processes. The first process was the chemical weathering of feldspar minerals containing silicon, such as NaAlSi3O8-CaAl2Si2O8. Reacting with CO2, these minerals produced acids that were washed into the ocean. Notably, calcium (Ca2+) and sodium (Na+) were washed away along with silicon. The second process was sulfate reduction. Unfortunately, this reaction did not produce any silicon. Instead, it produced bicarbonate (HCO3–). The abundance of bicarbonate, calcium, and sodium ions contributed to the alkalization of the ocean. The resulting “soda ocean” favored the dissolution of silicon (Muller et al., 2011). Its concentration increased up to 1000-2200 micromolars and remained higher than 650 micromolars up to the Jurassic period (145-201.2 Ma) (Maldonado et al., 2011) (Iqbal, 2021).
Having successfully occupied their niche for 400-500 million years, sponges were disturbed at the end of the late Cretaceous (65-96 Ma) (Sims et al., 2006). The diversification of diatoms, a type of green algae, endangered their peaceful existence. Needing silicon to grow their siliceous shells called frustules, diatoms developed an active pumping mechanism to siphon silicon out of their environment. As a result, silicon levels in the oceans decreased to an average of 10μM (Maldonado et al., 2011). Sponges were in danger as their essential resource was becoming scarcer and scarcer. Some evolved spongin and calcareous skeletons (Maldonado et al., 2020). However, 75% of extant sponges still have siliceous skeleton and consume 8.6 X 1010 and 7.3 x 1012 moles of silicon yearly (Maldonado et al., 2011).
Capturing Silicon
To begin building its skeleton, a sponge needs silicon. It needs to capture this element from the environment and concentrate it inside of its body. To this end, sponges developed a two-step transport system (Fig. 16). First, aquaglyceroporins 3 and 9 let silicon enter the epithelium cells of the sponge. Second, transporters resembling the arsenite-antimonite (arsB) and low-silicon (Lsi2) transporters actively transport silicon outside of the epithelial cell using the energy of a proton gradient (Schuldiner, 2014). The arsB and Lsi2 transporters underpin the proper function of the aquaglycoroporins because their active silicon efflux creates the gradient necessary for the passive silicon influx through the latter. From these findings, Maldonado et al. concluded that the active transporters must be responsible for the saturation of the sponge silicon capturing system in the range of 150-200μM. They also proposed that the promiscuity of these proteins barred the improvement of their silicon transport speeds (Maldonado et al., 2020).
Fig. 16. The two-step fixation of silicon by sponges. Dissolved silicon (DSi) enters the epithelial cells of the sponge by passive transport across aquaglyceroporins 3 and 9 (Aqua). Using the energy of a proton gradient, silicon is actively transported outside of the epithelial cells into the mesohyl, via arsenite-antimonite and low-silicon-like transporters (arsB). Later, sclerocytes absorb the silicon and deliver it to their silica deposition vesicle where the synthesis of a biosilica (BSi) spicule happens. (Maldonado et al., 2020).
On the other side of the epithelium, silicon enters a dense layer of extracellular matrix within the sponge – the mesohyl. In this environment, it triggers the differentiation of progenitor cells into sclerocytes (the cells that produce spicules) and promotes the synthesis of collagen to hold the newly made spicules (Muller et al., 2011).
Making Spicules
Once the raw material is inside of the sponge, the synthesis of the spicule begins. Sclerocytes absorb silicon containing molecules from the mesohyl and express silicatein, the enzyme that catalyses the formation of silica (Muller et al., 2011).
Silicatein reacts with orthosilicic acid, tetraethoxysilane, and organic oxysilanes leading to their polycondensation into silica. Figure 17 shows the species silicatein reacts with and Figure 18 shows the condensation reaction that leads to the synthesis of siloxane (Si-O-Si) bonds that characterize silica (Schroder et al., 2011).
Fig. 17. Substrates of silicatein. a) Orthosilicic acid in its monomeric and polymeric forms. b) Tetraethoxysilane. c) Bis(p-aminophenoxy)-dimethylsilane, an organic oxysilane. [a) Adapted from Schroder et al., 2011; b) Adapted from Wikipedia contributors; c) Adapted from National Center for Biotechnology Information].
Fig. 18. Mechanism leading to the formation of a siloxane bond between two orthosilicic acid molecules. (Schroder et al., 2011).
At its active site, silicatein has three amino acids that form its catalytic triad: asparagine, histidine, and serine (Fig. 19) (Muller et al., 2011). During catalysis, the oxygen atom of a hydroxyl group of serine attacks the silicon of the orthosilicic acid following an SN2 mechanism. At the same time, the hydrogen of the same hydroxyl group (which had formed a hydrogen bridge with histidine) attacks one of the hydroxyl groups of the substrate. This step results in the release of the hydroxyl group as a water molecule and the bonding of the substrate with the serine. In the next step, histidine and asparagine form hydrogen bridges with the substrate attached to the serine. The new position of the substrate-serine complex favors attacks by other orthosilicic acid molecules. Thus, leading to polycondensation (Schroder et al., 2011).
Fig. 19. Structure and activity of silicatein. a) Monomeric orthosilicic acid (Si(OH)4) in the catalytic pocket of silicatein surrounded by the catalytic amino acid triad in blue. b) Initial catalytic step. The oxygen of serine (Ser) attacks the silicon of the substrate, and the hydrogen of histidine (His) attacks one of its hydroxyl groups. c) Substrate-serine complex oriented for the next attack by hydrogen-bridges with histidine and asparagine (Asn) illustrated in yellow. [Adapted from Schroder et al., 2011].
When forming a spicule, having silicon and silicatein is not enough. The process needs to be coordinated to produce the right structure. It begins by the formation of the axial filament rich in silicatein within the sclerocyte. The filament serves as a scaffold for the subsequent deposition of multiple layers of silica. Significantly, silicatein is present both within the layers of silica and on the surface of the spicule contributing to its growth. When the first layer of silica is deposited, the spicule is ejected and grows in the mesohyl. Their silicatein binds to galectin molecules and arranges into strings parallel to the surface of the spicule. In later stages, the strings bind to collagen fibres, form a net and envelop the spicule (Muller et al., 2011).
While this mechanism of growth accounts for the diversity of spicule morphology, Mohri et al. identified an additional factor that contributes to it. Their transcriptomics analysis of the expression of silicatein in the different sclerocytes of freshwater demosponge Ephydatia fluviatilis showed surprising results. Six isoforms of silicatein, similar enzymes whose sequence is modified during production, exhibit different temporal activity in different sclerocytes. As a result, the cells produce different types of spicules (Fig. 20) (Mohri et al., 2008).
Fig. 20. E. fluviatilis has two types of sclerocytes. Megasclerocytes produce megascleres (top), and gemmosclerocytes produce gemmoscleres (bottom). The figure shows the development of spicules with the corresponding pattern of silicatein isoform expression with respect to time. The length of the red band to the right of each isoform reflects the duration of its expression, and the hue of the band, the relative amount of its mRNA in the cell. (Mohri et al., 2008).
Arranging the Skeleton
When all the spicules are made, the skeleton needs to be assembled like a puzzle. Unlike all other animals, sponges synthesize their skeletal elements away from their final position. Because of this unique trait, sponge cells need to cooperate to arrange the skeleton like a colony of termites would arrange their nest. The process is even more significant as it is the only known instance of cells cooperating to work with an inorganic material.
Nakayama et al. discovered the steps of this process in E. fluviatilis. Its cells divide the labour and iterate a sequence of “transport-pierce-raise up-cementation” steps (Fig. 21). At the beginning, a group of transport cells transports the spicule along the internal epithelium of the mesohyl. In a favorable location, the cells pierce the epithelium and embed half of the spicule in it. They also raise the spicule up. Basopinacocytes cement the dangling spicule at the base of the sponge with collagen fibers. Similarly, spicules can be added to already fixed ones by travelling along the upper surface of the epithelium. The iteration of these steps results in a pole-and-beam structure molded in accordance with the shape of the external environment (Nakayama et al., 2015).
Fig. 21. Assembly of the skeleton of E. fluviatilis through the “transport-pierce-raise up-cementation” process. (Nakayama et al., 2015).
Conclusion on Silica Biomineralization
In sum, high silicon levels characterized the environment in which sponges evolved 542-580 million years ago, and silicon became a key resource for these animals. As they developed silicon-fixing and silica-producing systems, sponges became able to build skeletons. Silicatein and collagen produced by multiple cooperating cells participated in the formation of larger and more complex skeletal structures. Despite being outcompeted by diatoms after 400-500 million years of thriving, sponges are still important players in oceanic silicon cycling.
Spongin: A Structural Protein with Superpowers
Another vital structural component of sea sponges is spongin.
Role and Composition of Spongin
Spongin provides the three-dimensional scaffold that supports sponge tissues in many species of Porifera, mainly in Demospongiae, the largest class of sponges, encompassing over 90% of the species in the phylum. Its 3D fibrous network (Fig.22) allows sponges to withstand dynamic marine currents and stresses while providing pores as entrances for feeding channels (Pozzolini et al., 2021).
Fig. 22. A. Photograph of a typical marine sponge skeleton, as what would be used as a commercial bath sponge B. Scanning electron microscope (SEM) micrograph showing the honeycomb-like microstructure of the marine spongin. (Domingues et al., 2021)
Produced by spongioblasts, spongin remains an enigmatic material for which the exact chemical structure has yet to be defined. While its classification as a collagen has been debated in literature, it would be indisputable to define it as a collagen-like biopolymer (Ehrlich et al., 2018). Indeed, like collagen, spongin consists of long chains of amino acids woven together in a triple-helical structure (Fig. 23) that provides both tensile strength and flexibility, and is predominately constituted of glycine, proline, and hydroxyproline amino acids (table 1) (Pozzolini et al., 2021).
Fig. 23. Triple-helical structure shared by collagen and spongin (Patel et al., 2022). The strands are intertwined, and, in spongin, bonded together by cross-linking of the strands (not shown).
Table 1. Amino acid composition of spongin filaments in I. oros and S. foetidus sponge samples compared with collagen from skin of codfish and commercial rat collagen (per 1000 residues) (Pozzolini et al., 2021). Notice the high quantities of glycine in all four samples, as well as the elevated content of hydroxylysine in the sponge samples.
Distinctive features of spongin include its significant halogen incorporation, notably of iodine and bromine, which cannot be found in terrestrial collagens and keratins (Żółtowska et al., 2021) (table 2). This halogenation helps explain the characteristic resistance these biopolymers have to traditional enzymatic treatment by amylases, lysozymes, collagenases and other proteases. Nonetheless, in their natural environment, several bacteria are able to enzymatically process spongin, but the isolation of special “sponginases” has yet to be successful (Ehrlich et al., 2018).
Table 2. Chemical composition of spongin (Ehrlich et al., 2018). Notice the presence of iodine and bromine, which distinguishes it from other collagens.
Engineering applications of Spongin
Spongin has been harvested, sold, and used for millennia for its absorption properties in applications related to cleaning, to the point where the application has been intertwined with the namesake of the sponge, even after cleaning tools transitioned to being made of synthetic materials. While the term spongin was only coined by Georg Städeler in 1859, the material was already valued in the medical world for its softness, high compressive strength, ability to retain shape, and high sorption rates, as well as in the pharmaceutical world for its iodine content as early as in the 18th century (Ehrlich et al., 2018) (Sipkema et al., 2005).
Nowadays, spongin is an intriguing material to use in water purification processes.
To Remove Mercury in Water Remediation
The presence of heavy metals in water is a threat to all forms of life but can be an especially concerning problem for apex predators, as these contaminants bio-accumulate in the food chain. Mercury (Hg) is classified as the third most dangerous substance in the Agency for Toxic Substances and Disease Registry (ATSDR)’s substance priority list, and can cause harm to the human immune system, kidneys, and lungs if present in the body. Therefore, remediating industrial and domestic wastewater containing Hg is a priority.
Spongin is a promising material that can serve as a sustainable, economical, and eco-friendly option for mercury remediation. Indeed, in their natural environments, sponges serve as biomonitors for several contaminants, as they sorb a multitude of contaminants in their environment by virtue of filter feeding. Consequently, it is possible to harness the chemical properties and lattice structure of spongin to create a natural water treatment system that can be easily separated from the treated water without requiring time-consuming processes such as filtration or centrifugation (Domingues et al., 2021). Moreover, marine sponges can easily be cheaply obtained in large quantities, as sponge farms are already a well-established industry.
To extend the use of spongin materials to harsh conditions such as stirred media, it is possible to increase its stiffness by heat-treating it at 180˚C, as it is the highest temperature at which the adsorbed water is lost, but also not hot enough for the degradation of the organic phase of the protein can occur at 200˚C (Fig. 24). This treatment results in a significant volume shrinkage, with the volume of open pore space going down from 85.2% to 51.6% (Domingues et al., 2021).
Fig. 24. Photograph and CT scan of a sample of MS before and after heat-treatment at 180˚C (Domingues et al., 2021). Notice the reduced open pore space in the treated sample.
Fourier-transform infrared spectroscopy (FTIR) can identify functional groups on the spongin sorbent’s surface, aiding in mapping its chemical structure (Fig. 25). The broad peaks in the 3600–3100 cm−1 interval indicate the stretching vibrations of N−H and O−H, while stretching vibrations of −CH, −CH2 and −CH₃ can be ascribed to the peaks in the 2950–2850 cm−1 range, and those of C=O, CO−NH, C−OH, C−O−C, and O=C−N are associated with the peaks at ∼1630 cm−1 , ∼1520 cm−1 , ∼1030 cm−1 , ∼1230 cm−1 , and ∼550 cm−1 , respectively (Domingues et al., 2021). More peaks associated with ring-bearing amino acids, deformation vibration of −CH₃, and others are shown in figure 4.
Fig. 25. FTIR spectra of untreated, heat-treated at 180 ˚C and post-contact with Hg solution MS samples (Domingues et al., 2021).
Studying mercury sorption rates (R%), we find that in an ultrapure water solution at 50 and 500 μg/L of Hg, the sorption rates are both high (91 and 94%, respectively), meaning that there is little dependence between the mercury concentration and the sorption rate. Furthermore, the effect of the spongin dosage with regards to sorption rate is also insignificant (Fig. 26B). However, as seen in figure 4C, the heat-treated spongin sample has significantly lower sorption rates than that of the untreated sample (Domingues et al., 2021).
Fig. 26. A) Effect of initial Hg concentration on sorption efficiency B) Effect of increasing MS dosage on the sorption efficiency. C) Effect of MS thermal treatment at 180˚C on sorption efficiency (Domingues et al., 2021). The bottom scale indicates time (h). Initial Hg concentration in B and C is 50 μg/L.
While the water initially at 50 μg/L Hg treated with the spongin sorption agent had residual mercury concentrations of around 5 μg/L, above the Canadian maximum acceptable concentration in drinking water of 1 μg/L, this treatment remains promising, as it is completely natural, easy, and economically viable. Moreover, no chemical functionalization has been applied to the spongin. Thus, with adequate additions to the surface chemistry of the biopolymer, it is possible to increase the efficiency of the treatment to a level where it can be commercially implemented.
As a Scaffold for Catalysis
Spongin can be thermally stable at high temperatures. Indeed, there are hydroxyproline residues capable of forming hydrogen bonds between the strands of this biopolymer as well as a high content of hydroxylysine (Fig. 22), which is known to be involved in crosslinking reactions, and the more crosslinks there are, the more stable it can be at high temperatures (Pozzolini et al., 2021). This stability also allows it to be carbonized while retaining its structural integrity and porous nature, conferring it various industrial applications (Żółtowska et al., 2021).
Carbonized spongin-based scaffolds have demonstrated significant efficacy in supporting nickel-based catalysts, especially in the context of environmental remediation. Research conducted by Żółtowska et al. (Żółtowska et al., 2021) involved the modification of spongin-based scaffolds with nickel compounds through a sorption-reduction technique, resulting in the formation of three distinct NiO/Ni(OH)₂/Ni composites. These composites, originating from the carbonized spongin scaffolds, exhibited exceptional catalytic performance in both oxidation and reduction reactions.
The fibrous three-dimensional structure of the spongin-derived scaffolds is pivotal to the catalytic processes. The distinctive architecture of these biocarbons allows for effective dispersion of the nickel phase and promotes interactions with reactants. This structural benefit, in conjunction with the catalytic characteristics of the nickel phase, establishes a highly efficient platform for catalytic applications. Consequently, these composites are adept at catalyzing oxidation reactions of phenolic compounds, such as methylchlorophenoxypropionic acid (MCPP) and 4-chlorophenoxyacetic acid (4-CPA), achieving oxidation efficiencies of up to 99%, as well as facilitating the reduction of 4-nitrophenol (Żółtowska et al., 2021).
The incorporation of heteroatoms such as bromine, iron, and nitrogen within the spongin-based scaffolds enhances the functionalization of the resultant biocarbons. These heteroatoms generate a variety of surface functional groups, which improve the adsorption and catalytic capabilities of the composite materials (Norman et al., 2018). Furthermore, the carbonized spongin-based supports exhibit remarkable chemical and thermal stability, rendering them suitable for catalytic applications across diverse environmental conditions.
Conclusions
Sponges have developed numerous chemical design solutions to thrive in their aquatic environment. This includes multiple feeding techniques and partnerships between other organisms. Chemical defense mechanisms allow the sponges to survive in crowded environments or areas with mobile predators. Sponges can produce chemicals to prevent the settlement of other organisms, they can create chemical shields around their bodies, and they have developed mechanisms to avoid self-toxicity. Additionally, even without the silicon density they once had, sponges are designed to collect silicon from the environment to form their spicules and make their skeleton. Sponges are also supported by spongin which is a material that provides support to sponges against the environmental forces. All in all, sponges are chemically diverse creatures that have developed brilliant methods to survive in their environment, which humans can learn from utilize in the form of pharmaceuticals and biomimetic structures.
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