Leveraging Functional Genomics and Engineering Approaches to Uncover the Molecular Mechanisms of Cnidarian–Dinoflagellate Symbiosis and Broaden Biotechnological Applications

Abstract

Functional genomics is a powerful approach for uncovering molecular mechanisms underlying complex biological processes by linking genetic changes to observable phenotypes. In the context of algal symbiosis, this framework offers significant potential for advancing our understanding of the molecular interactions between marine dinoflagellates and their cnidarian hosts, such as corals—organisms that are foundational to marine ecosystems and biodiversity. As coral bleaching and reef degradation intensify due to environmental stressors, novel strategies are urgently needed to enhance the resilience of these symbiotic partnerships. This opinion piece explores emerging directions in functional genomics as applied to coral–algal symbiosis, with a focus on uncovering the molecular pathways that govern photosynthesis and stress tolerance. We discuss the challenges and opportunities in applying functional genomics to support coral health, improve ecosystem resilience, and inform biotechnological applications in agriculture and medicine. Together, these insights posit the potential for engineered symbioses as a needed focus in mitigating biodiversity loss and supporting sustainable ecosystem management in the face of accelerating environmental change.

1. Introduction

Endosymbiotic dinoflagellates in the family Symbiodiniaceae play a foundational role in the health and resilience of coral reef ecosystems [1] (Figure 1). In symbiosis with cnidarian hosts—including corals, sea anemones, and jellyfish—these algae fuel reef productivity by supplying photosynthates (e.g., sugars, amino acids, lipids) that support host metabolism and calcification, in exchange for protection and inorganic nutrients [2,3]. This metabolic exchange is central to the stability of cnidarian–Symbiodiniaceae symbiosis and supports the productivity and ecological function of coral reef ecosystems, which are among the most biologically diverse habitats in the ocean [4,5,6,7]. However, this symbiosis is increasingly threatened by environmental stressors such as ocean warming and acidification, leading to coral bleaching, energetic deficits, and increased susceptibility to disease [8,9,10,11,12,13]. Beyond their ecological role, endosymbiotic dinoflagellates are garnering interest in biotechnology for their ability to produce diverse bioactive metabolites with potential applications in pharmaceuticals, nutraceuticals, and other industries [14,15].

Functional Genomics to Decipher and Engineer Marine Symbioses

In this context, functional genomics offers a powerful framework for understanding biological processes and linking genotype to phenotype, thereby advancing insights into gene function [16]. For this purpose, functional genomics often involves disrupting or modifying an organism’s genome to observe the resulting phenotypic changes [17,18]. Techniques such as CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-based genome editing, UV mutagenesis, and RNA interference (RNAi) have been instrumental in elucidating pathways underlying processes as diverse as osteoarthritis progression and chlorophyll production in marine algae [19,20,21,22,23]. In marine systems, functional genomics provides unique opportunities to unravel novel and unknown biosynthetic pathways and investigate mutualistic relationships, such as those between algae and cnidarians like corals [24,25]. This opinion piece highlights the potential of functional genomics to guide the engineering of symbiotic partners to enhance theirresilience against coral bleaching and mitigate biodiversity loss. These advances can also be applied to agricultural improvements in terms of photosynthetic efficiency and environmental stress tolerance [26,27].

2. Photosynthesis in Marine Algal Symbiosis

The study of photosynthesis within cnidarian–algal symbiosis is critical in understanding how environmental changes affect marine ecosystems [28]. Functional genomics has been instrumental in advancing this understanding, as demonstrated by the application of UV mutagenesis to Symbiodiniaceae algae, which provides a promising platform to investigate symbiont gene function and its role in the partnership [29]. Recent experiments using photosynthetically impaired algal mutants revealed that active photosynthesis is not required for the establishment of symbiosis, although it appears to be critical for supporting a healthy symbiont population [29,30,31]. These findings underscore the complexity of host–symbiont interactions and highlight the value of functional genomics in uncovering molecular mechanisms underlying symbiotic resilience.

Cnidarian–algal symbiosis establishment is characterized by three stages: infection, proliferation, and maintenance. Infection marks the initiation of cnidarian–algal symbiosis where cnidarian hosts acquire algal cells from the environment into their gastrodermis [2,29,32]. During infection, symbiont selection is thought to involve a winnowing process, potentially mediated by algal properties and host recognition events [33]. Upon successful infection, symbiont algae (and to a degree, host cells) undergo a period of rapid proliferation in hospite [34]. Maintenance occurs after infection and proliferation, when a steady-state algal population within the host is achieved under given environmental conditions [2].

2.1. The Establishment of Cnidarian–Symbiodiniaceae Symbiosis Is Independent of Algal Photosynthesis

One characteristic trait of Symbiodiniaceae algae—photosynthesis—is thought to be a factor in symbiosis establishment. Studies with the Breviolum minutum strain SSB01 show that culturing algae in glucose-containing media under continuous light reduces their photosynthetic function and ability to infect sea anemone hosts [35]. However, these conditions also alter other physiological traits, such as cell surface complexity, complicating efforts to establish causality. To address this knowledge gap, Jinkerson et al., 2022 further investigated the role of photosynthesis during infection, proliferation, and maintenance stages of cnidarian–Symbiodiniaceae symbiosis by using darkness, a chemical inhibitor of photosynthesis (DCMU), and photosynthetic B. minutum mutants [29]. Cnidarian model systems, including with Exaiptasia pallida (Aiptasia), coral Acropora tenuis, and jellyfish Cassiopea xamachana, were used as hosts with B. minutum SSB01 and four additional symbiont species—Symbiodinium linucheae, Cladocopium goreaui, Durusdinium trenchii, and Symbiodinium necro appetens—to evaluate symbiotic establishment and account for species-specific host–symbiont interactions. Symbiont infection, proliferation, and maintenance of algal cells were evaluated and quantified using fluorescent imaging, revealing that symbiosis establishment occurs in continuous darkness and in the presence of DCMU. Following these initial experiments, the study employs functional genomics to generate B. minutum mutants and evaluate whether symbiosis establishment between cnidarian hosts and algal symbionts can occur independently of photosynthesis. Due to the lack of available transformation methods for Symbiodiniaceae, the study employs UV mutagenesis to introduce genetic disruptions. Mutants, initially screened for altered coloration, are followed by secondary screening to identify photoautotrophic growth deficiencies [36]. This approach yields six symbiont mutants, including LIGHT BROWN 1 (lbr1), ORANGE 1 (ora1), and YELLOW 7-9,12 (yel7, yel8, yel9, yel12). These mutants are then tested for their ability to establish, proliferate, and maintain symbiosis across three host systems [29].

The findings of the study by Jinkerson et al., 2022 reveals that the establishment of this symbiosis is specific to the cnidarian host and can occur independently of the photosynthetic ability of symbiont algae [29]. Specifically, their assessment across 31 host–symbiont pairs shows that photosynthesis is not necessary for Symbiodiniaceae to infect Aiptasia, coral, or Cassiopea, nor is it required for symbiont maintenance within cnidarian hosts. In particular, B. minutum was observed to be present in Aiptasia tissue for 180 days post infection in continuous darkness. However, the extent of symbiont proliferation in the absence of photosynthesis varied by host. While Acropora polyps supported algal proliferation without photosynthesis, both Aiptasia and Cassiopea showed limited symbiont growth under similar conditions [29] (Figure 2). This plasticity points to alternative metabolic interactions that may support symbiosis during stress or developmental transitions, such as in early life stages. For instance, juvenile Acropora polyps were able to support algal proliferation even in the absence of photosynthesis, hinting at life-stage-specific nutrient provisioning strategies.

2.2. Other Molecular and Environmental Modulators of Symbiosis Establishment

These observations raise important questions about the alternative molecular and cellular mechanisms that facilitate symbiosis in the absence of photosynthesis. For example, the ability of cnidarian–algal associations to initiate independently of photosynthesis highlights the possible involvement of host immune modulation and recognition pathways [32,37]. In the Hydra–Chlorella model, for instance, microbe-associated molecular pattern (MAMP) recognition has been shown to mediate immune tolerance and help maintain symbiosis, suggesting that immune signaling may be central to initiating stable host–symbiont interactions [38]. Complementing these molecular insights, environmental conditions—including light, temperature, and pH—further modulate the physiological dynamics of the symbiotic relationship [39,40,41]. Light intensity, in particular, has a profound effect on dinoflagellate physiology, regulating growth, pigment composition, and stress responses. Transcriptomic analyses reveal that changes in environmental light conditions significantly alter gene expression in B. minutum, including genes involved in light harvesting, cell adhesion, and nutrient transport [42]. In Durusdinium glynnii, high light exposure induces the accumulation of peridinin, a photoprotective carotenoid that shields chlorophyll from photo-oxidation, suggesting an adaptive response to elevated irradiance [43]. Symbionts in the tropical hydroid Myrionema amboinense further demonstrate the ability to acclimate to low-light environments [44]. Beyond light, environmental stressors such as ocean acidification and pollutants can lower pH, often acting synergistically with warming and irradiance to trigger bleaching responses [45,46]. These findings underscore the multifactorial nature of symbiotic regulation and point to immune signaling, environmental sensing, and metabolic plasticity as key targets for engineering more resilient host–symbiont partnerships.

3. Research Beyond the Role of Photosynthesis Within Cnidarian–Algal Symbiosis

While photosynthesis is not essential for some steps in cnidarian–algal symbiosis, it remains a critical energetic pathway that ultimately supports nearly all life on Earth [4,47]. Beyond its ecological importance, photosynthesis also supplies and sustains human agriculture, yet its energy conversion efficiency in crop plants is estimated to be as low as 1% [48,49]. Therefore, improving photosynthetic efficiency is vital to meet growing agricultural demands; strategies include widening the usable light spectrum, enhancing light capture, and optimizing carbon fixation [50,51].

3.1. Identification of Chlorophyll c Biosynthesis Pathway in Dinoflagellates

In particular, researchers have explored strategies to enhance photosynthetic efficiency in crops by incorporating foreign chlorophyll pigments. One such pigment, chlorophylls c (Chls c), is predominantly found in marine algae such as diatoms, brown algae, dinoflagellates, and many other lineages of eukaryotic algae, where it captures blue-green wavelengths (447–452 nm) critical for photosynthesis in aquatic environments [52,53,54]. The absence of Chls c in terrestrial plants implies distinct ecological and evolutionary adaptations in marine algal species and represents the importance of understanding its biosynthetic pathways [55,56]. Elucidating the biological production pathway of critical chemicals, like chlorophyll c, is urgently needed for leveraging these pathways to optimize agricultural yields [57].

As such, the biosynthesis of Chls c in dinoflagellate Symbiodiniaceae has been investigated through the analysis of B. minutum photosynthesis mutants [20]. Among them, the mutant lbr1 exhibits less light absorbance from its typical blue–green spectrum. Upon further investigation with ultra-high-performance liquid chromatography–high-resolution mass spectrometry (UHPLC-HRMS), they discover that the lbr1 mutant, in particular, was unable to produce Chls c. Sequencing of its transcriptome further reveals multiple mutations, including a frameshift mutation in gene s6_3623. The study identifies the s6_3623 gene as the causal gene for Chls c production by heterologous expression of s6_3623 in Nicotiana benthamiana. Determining that this gene codes for an enzyme, identified and named “BmCHLCS (CHLOROPHYLL C SYNTHASE),” the study confirms its vital role in algal Chls c biosynthesis through the heterologous production of Chls c in N. benthamiana [20].

To address the initially limited production of Chls c in N. benthamiana extracts, the researchers use a dual-intervention approach consisting of the usage of an δ-Aminolevulinic acid (ALA) substrate followed by dark incubation. The resulting increase in Chls c production in N. benthamiana enables mass spectrometry detection of peaks corresponding to Chls c₁ and c_2., confirming the light independence of BmCHLCS. In order to determine the catalytic domain responsible for Chls c production, the study evaluates BmCHCLS function through assessment of the chlorophyll a/b binding and 2-oxoglutarate-Fe(II) dioxygenase (2OGD) domains. Subsequent assessment of BmCHCLS function by selective deletion of either of the two domains reveals that 2OGD is the catalytic domain as only 2OGD deletion affects the production of Chls c, resulting in no detectable levels of chlorophyll c₁ and c₂ in the leaves of N. benthamiana [20]. Enzymes with 2OGD dioxygenase domains are known to biosynthesize a wide range of metabolites in plants, including hormones, alkaloids, and other natural products, with most characterized pathways localized to the cytosol or mitochondria [58,59,60]. This study identifies a 2OGD-dependant pathway in the chloroplast, expanding the known sites for 2OGD-associated biosynthetic reactions and illuminating new opportunities for chloroplast engineering. Further investigation into BmCHLCS reveals that a bipartite signal and transit peptide sequence directs the enzyme to the chloroplast, enabling its function in Chls c biosynthesis. Interestingly, the chlorophyll a/b binding domain, while not essential for Chls c biosynthesis, appears to facilitate localization of BmCHLC in the chloroplast, highlighting a unique mechanism with potential implications for the enzyme’s structure and function [20].

A comprehensive analysis of CHLCS distribution reveals its presence in peridinin-containing dinoflagellates, while all chlorophyll c-dependent algal lineages encode 2OGD-containing proteins (CHLCs), forming a strongly supported monophyletic group in phylogenetic analyses. However, a subset of Ochrophyta lacks CHLCS or chlorophyll c homologs despite their reliance on chlorophyll c, suggesting the presence of alternative enzyme(s) responsible for chlorophyll c biosynthesis [20]. Investigating these pathways offers an exciting opportunity to uncover novel enzymes, shedding light on the evolution of pigment biosynthesis in ochrophytes and their adaptation to diverse ecological niches [61,62,63].

3.2. From Dinoflagellates to Plants for Enhanced Light Capture and Artificial Photosynthesis

Overall, the identification of the gene responsible for chlorophyll c production represents a significant milestone, given the inherent challenges of functional genomics in dinoflagellates. This study marks the first successful heterologous production of chlorophyll c in plants, opening the door to engineering terrestrial plants with expanded pigment diversity and enhanced light-harvesting capacity. As efforts intensify to develop crops with improved resilience to climate stress and more efficient solar energy capture, biomimetic strategies inspired by photosynthetic systems are gaining momentum [64]. Recent studies demonstrate how bioinspired designs can enhance photovoltaic performance while reducing heat output [65]. Parallel advances in artificial photosynthesis have demonstrated efficient CO₂ fixation and solar-to-food energy conversion, enabling the production of concentrated acetate streams that can support heterotrophic growth of various organisms—including photosynthetic algae—without reliance on natural photosynthesis [66]. Together, these innovations point to new frontiers in sustainable agriculture and controlled-environment food production, with the potential to surpass the efficiency of traditional photosynthetic systems.

4. Marine Pharmacology: A Sustainable Frontier for Therapeutic Discovery

Beyond just the use of functional genomics to uncover biosynthetic pathways in plant photosynthetic pigments, these strategies offer immense potential in marine systems, where they can illuminate the molecular mechanisms underlying the production of bioactive compounds [67]. Marine pharmacology represents a sustainable and innovative frontier for therapeutic discovery, bridging ecological research with clinical advancement. By exploring the vast chemical diversity of marine organisms—such as algae, cyanobacteria, sponges, and corals—this field seeks to identify novel bioactive compounds with applications in treating cancer, infection, inflammation, and neurological disorders [68,69,70]. These compounds, often shaped by evolutionary pressures in complex symbiotic or competitive marine ecosystems, offer unique structural scaffolds and mechanisms of action that are rarely found in terrestrial organisms [71,72]. To date, over 13,000 molecules derived from marine species have been identified, with approximately 3000 exhibiting significant pharmacological activity [73,74]. These compounds, primarily secondary metabolites, are not directly involved in essential biological processes such as growth or reproduction but possess immense therapeutic potential. They could offer solutions to pressing medical challenges, including cancer; bacterial, viral, parasitic, and fungal infections; and applications in bioregenerative medicine [75,76,77].

Bridging Gaps in Biosynthetic Mechanisms Is Critical for Novel Therapeutic Development

While the therapeutic potential of marine-derived compounds spans numerous fields, gastrointestinal research presents particularly promising and readily feasible translational opportunities [78,79,80]. Advances in gut microbiome modulation—such as microbial fecal implants and supplements—could mitigate current challenges, including poorly characterized disease mechanisms, reliance on complex animal models with limited clinical relevance, immune rejection of treatments, and uncertainties surrounding long-term outcomes [81,82,83]. Chronic and acute inflammatory gastrointestinal disorders, such as ulcerative colitis, diverticulitis, pancreatitis, and irritable bowel syndrome, are often linked to microbial imbalances or disruptions in microbe–host homeostasis [84,85]. Insights into these conditions could be gained by applying functional systems biology approaches that are informed by analogies to algal symbiosis and biosynthesis, facilitating improved diagnosis, prognosis, and therapeutic outcomes [14,86].

Despite its vast potential, marine pharmacology faces significant challenges. Comprehensive reviews by Malve et al., 2016 and Mayer et al., 2019 highlight the therapeutic promise of marine-derived pharmaceuticals and their capacity to expand the repertoire of available therapeutic agents [73,74]. However, only a small fraction of marine natural products and their derivatives have been approved or are undergoing clinical testing by the U.S. Food and Drug Administration (USFDA) [87,88,89,90]. This slow progress stems from gaps in understanding the biosynthetic pathways of these compounds, limiting the efficiency and sustainability of their extraction and formulation [15]. Addressing these obstacles through functional genomics and other advanced techniques will be critical for unlocking the full potential of marine systems in drug discovery and development.

5. Advancing Genomic Research in Marine Symbiosis and Beyond

Understanding marine symbiosis, particularly in cnidarian–algal interactions, is pivotal for ecological and biotechnological progress. This symbiosis establishment is impacted by a complex interplay of environmental factors such as light, temperature, salinity, and nutrient availability. Investigating the molecular and cellular mechanisms underlying algal infection into host gastrodermal cells, including processes like cell-surface recognition and symbiosome formation, remains an essential frontier. Unraveling these processes will provide crucial insights into the specificity and stability of symbiotic relationships, with broad implications for ecosystem resilience and health (Figure 3).

5.1. Limitations in Genetic Engineering Tools for Studying Symbiodiniaceae Algae

Genomic and genetic research in Symbiodiniaceae species also represents a critical avenue for future work. However, challenges posed by their unique genomic organization, including permanently condensed chromosomes and lack of traditional transcriptional regulation, limit the application of conventional genetic tools like CRISPR-Cas9 [91,92,93]. Expanding these efforts to sequence a broader range of Symbiodiniaceae genomes across diverse species is essential for capturing the genetic diversity and evolutionary adaptations within this ecologically important family [94,95,96]. Such diversity will provide valuable insights into species-specific traits, host compatibility, and environmental resilience [97]. Despite the importance of functional genomic studies, progress is limited by the complexity of genetic manipulation, which contrasts with the simpler methods used in bacterial, viral, or algal systems [98,99,100,101,102]. Promising advancements in cell-wall deconstruction have shown potential for improving transformation efficiency, offering a path forward for developing robust genetic manipulation methods [103]. Further inquiry into novel genomic approaches is vital for overcoming these barriers. Ultimately, integrating functional genomics with metabolomics and proteomics will help elucidate the molecular mechanisms of symbiosis and provide a framework for addressing broader ecological and biotechnological challenges [104,105].

5.2. Conclusions and Outlook

These insights hold broad applications beyond symbiosis research. Marine pharmacology, for example, highlights the therapeutic potential of marine bioactive compounds, which could address critical medical challenges. Specifically, understanding the biosynthetic pathways of these compounds through functional genomics could enhance sustainable drug development. Similarly, advancements in photosynthetic biology, such as research on chlorophyll c synthase in marine algae, offer pathways to improve photosynthetic efficiency and resilience in terrestrial crops. Integrating genomics with metabolomics and proteomics enables the discovery of the molecular mechanisms underlying symbiosis and photosynthesis, fostering innovations in sustainable agriculture, climate adaptation, and biomedicine. Robust genomic resources and methodologies tailored to Symbiodiniaceae are essential not only for unraveling the complexities of marine symbiosis, but also for leveraging marine systems to address global challenges in health, agriculture, and environmental sustainability.