Associated bacteria affect sexual reproduction by altering gene expression and metabolic processes in a biofilm inhabiting diatom

Diatoms are unicellular algae with a fundamental role in global biogeochemical cycles as major primary producers at the base of aquatic food webs. In recent years, chemical communication between diatoms and associated bacteria has emerged as a key factor in diatom ecology, spurred by conceptual and technological advancements to study the mechanisms underlying these interactions. Here, we use a combination of physiological, transcriptomic, and metabolomic approaches to study the influence of naturally coexisting bacteria, Maribacter sp. and Roseovarius sp., on the sexual reproduction of the biofilm inhabiting marine pennate diatom Seminavis robusta. While Maribacter sp. severely reduces the reproductive success of S. robusta cultures, Roseovarius sp. slightly enhances it. Contrary to our expectation, we demonstrate that the effect of the bacterial exudates is not caused by altered cell-cycle regulation prior to the switch to meiosis. Instead, Maribacter sp. exudates cause a reduced production of diproline, the sexual attraction pheromone of S. robusta. Transcriptomic analyses show that this is likely an indirect consequence of altered intracellular metabolic fluxes in the diatom, especially those related to amino acid biosynthesis, oxidative stress response, and biosynthesis of defense molecules. This study provides the first insights into the influence of bacteria on diatom sexual reproduction and adds a new dimension to the complexity of a still understudied phenomenon in natural diatom populations

Distinctive growth and transcriptional changes of the diatom Seminavis robusta in response to quorum sensing related compounds

In aquatic habitats, diatoms are frequently found in association with Proteobacteria, many members of which employ cell-to-cell communication via N-acyl homoserine lactones (AHLs). It has been suggested that diatoms could distinguish between beneficial and algicidal bacteria in their surroundings by sensing AHLs. Although some microalgae can interfere with AHL signaling, e.g., by releasing AHL mimics or degrading them, molecular responses to AHLs in microalgae are still unclear. Therefore, we tested the effects of short-chained AHLs, i.e., N-hexanoyl homoserine lactone (C6-HSL), N-3-hydroxyhexanoyl homoserine lactone (OH-C6-HSL), and N-3-oxohexanoyl homoserine lactone (oxo-C6-HSL) and long-chained AHLs, i.e., N-tetradecanoyl homoserine lactone (C14-HSL), N-3-hydroxytetradecanoyl homoserine lactone (OH-C14-HSL), and N-3-oxotetradecanoyl homoserine lactone (oxo-C14-HSL), on growth of the benthic diatom Seminavis robusta. All tested short-chained AHLs did not affect diatom growth, while long-chained AHLs promoted (C14-HSL) or inhibited (OH-C14-HSL and oxo-C14-HSL) growth. To investigate the physiological effects of these long-chained AHLs in more detail, an RNA-seq experiment was performed during which S. robusta was treated with the growth-promoting C14-HSL and the growth-inhibiting oxo-C14-HSL. One tetramic acid was also tested (TA14), a structural rearrangement product of oxo-C14-HSL, which also induced growth inhibition in S. robusta. After 3 days of treatment, analysis revealed that 3,410 genes were differentially expressed in response to at least one of the compounds. In the treatment with the growth-promoting C14-HSL many genes involved in intracellular signaling were upregulated. On the other hand, exposure to growth-inhibiting oxo-C14-HSL and TA14 triggered a switch in lipid metabolism towards increased fatty acid degradation. In addition, oxo-C14-HSL led to downregulation of cell cycle genes, which is in agreement with the stagnation of cell growth in this treatment. Combined, our results indicate that bacterial signaling molecules with high structural similarity induce contrasting physiological responses in S. robusta

Mating type specific transcriptomic response to sex inducing pheromone in the pennate diatom Seminavis robusta

Sexual reproduction is a fundamental phase in the life cycle of most diatoms. Despite its role as a source of genetic variation, it is rarely reported in natural circumstances and its molecular foundations remain largely unknown. Here, we integrate independent transcriptomic datasets to prioritize genes responding to sex inducing pheromones (SIPs) in the pennate diatomSeminavis robusta. We observe marked gene expression changes associated with SIP treatment in both mating types, including an inhibition of S phase progression, chloroplast division, mitosis, and cell wall formation. Meanwhile, meiotic genes are upregulated in response to SIP, including a sexually induced diatom specific cyclin. Our data further suggest an important role for reactive oxygen species, energy metabolism, and cGMP signaling during the early stages of sexual reproduction. In addition, we identify several genes with a mating type specific response to SIP, and link their expression pattern with physiological specialization, such as the production of the attraction pheromone diproline in mating type - (MT-) and mate-searching behavior in mating type + (MT+). Combined, our results provide a model for early sexual reproduction in pennate diatoms and significantly expand the suite of target genes to detect sexual reproduction events in natural diatom populations

Scaling the invisible wall : molecular acclimation of a salinity‐tolerant diatom to freshwater

In aquatic ecosystems, marine and freshwater environments are separated by steep salinity gradients. The osmotic stress induced by this 'invisible wall' forms an insurmountable barrier for many aquatic lifeforms, including bacteria, algae and animals. Because the osmotic differences when transiting a salinity divide are so hard to overcome, most species have adapted exclusively to a marine or a freshwater lifestyle. A major consequence of this physiological specialization into marine and freshwater organisms is that transitions are relatively rare, impeding regular contact and colonization. While some animals use specialized organs or behaviour to cope with unfavourable salinity levels, unicellular algae such as diatoms are completely dependent on cellular mechanisms to mitigate salinity stress. In this issue of Molecular Ecology, Downey and colleagues investigate the transcriptomic response of a salinity-tolerant diatom to a shock treatment with freshwater (Molecular Ecology, 2023). Through frequent sampling and integration of existing RNA sequencing data, a fine-grained model of the acclimation to hypo-osmotic stress emerges. Deciphering the pathways that drive the acute and long-term acclimation to freshwater has major implications for diatom ecology, diversification and resilience to global change

Transcriptional responses to salinity‐induced changes in cell wall morphology of the euryhaline diatom Pleurosira laevis

Diatoms are unicellular algae with morphologically diverse silica cell walls, which are called frustules. The mechanism of frustule morphogenesis has attracted attention in biology and nanomaterials engineering. However, the genetic regulation of the morphology remains unclear. We therefore used transcriptome sequencing to search for genes involved in frustule morphology in the centric diatom Pleurosira laevis, which exhibits morphological plasticity between flat and domed valve faces in salinity 2 and 7, respectively. We observed differential expression of transposable elements (TEs) and transporters, likely due to osmotic response. Up-regulation of mechanosensitive ion channels and down-regulation of Ca2+-ATPases in cells with flat valves suggested that cytosolic Ca2+ levels were changed between the morphologies. Calcium signaling could be a mechanism for detecting osmotic pressure changes and triggering morphological shifts. We also observed an up-regulation of ARPC1 and annexin, involved in the regulation of actin filament dynamics known to affect frustule morphology, as well as the up-regulation of genes encoding frustule-related proteins such as BacSETs and frustulin. Taken together, we propose a model in which salinity-induced morphogenetic changes are driven by upstream responses, such as the regulation of cytosolic Ca2+ levels, and downstream responses, such as Ca2+-dependent regulation of actin dynamics and frustule-related proteins. This study highlights the sensitivity of euryhaline diatoms to environmental salinity and the role of active cellular processes in controlling gross valve morphology under different osmotic pressures

Cellular hallmarks and regulation of the diatom cell cycle

Diatoms are one of the oldest experimental models for studying the mitotic cell cycle, with microscopic descriptions of cell division dating back to the nineteenth century. In recent years, the advent of genetic and genomic tools has improved our understanding of the mechanisms driving cell cycle progression in diatoms. Diatom species thrive in almost all aquatic habitats and several of them form blooms under specific environmental conditions. In order to optimize their growth rate to the prevailing conditions, species-specific cell cycle checkpoints have evolved that integrate cues such as light, nutrients, and sex pheromones. This chapter reviews the structural events occurring during each cell cycle stage, focusing on organelle division, the unique mitotic spindle of diatoms, and the different steps of mitosis. The conservation of the core cell cycle components in diatom genomes is briefly explored. External conditions that activate the G1/S and G2/M checkpoints of the interphase are discussed, with special attention to the light-dependent G1 phase checkpoint in P. tricornutum, which is currently the best characterized regulatory cell cycle pathway in diatoms. The chapter concludes with an outlook on how novel technologies can contribute to solving the specificities of the diatom cell cycle at a molecular level

Life cycle regulation

Diatom life cycles are unusual among microalgae by being diplontic with a long diploid vegetative phase and a short-lived haploid phase (gametes). Life cycle progression in diatoms is controlled by the cell size reduction-restitution cycle and is intimately linked to their peculiar mode of cell division and siliceous cell wall. Sexual reproduction is primarily cell-size dependent although environmental cues may be needed to trigger gametogenesis in centric diatoms. Although population genetic data suggest sexual reproduction to occur in most species and meiotic genes are widely conserved among diatoms, sexual events are seldom observed in nature. Recent laboratory studies have started to unveil complex pheromone signaling cascades during sexual reproduction in pennate diatoms. Likewise, significant progress has been made in the identification of mating type determination mechanisms in heterothallic species, where several conserved, but as yet functionally uncharacterized, genes involved in sexual reproduction have been identified. While many aspects of diatom life cycle regulation remain to be discovered, the recent development of new model species allowing genetic modification and the rapidly increasing genomic and transcriptomic resources hold much promise for advanced understanding of this key process