Mechanisms regulating the intracellular trafficking and release of CLN5 and CTSD.

Ceroid lipofuscinosis neuronal 5 (CLN5) and cathepsin D (CTSD) are soluble lysosomal enzymes that also localize extracellularly. In humans, homozygous mutations in CLN5 and CTSD cause CLN5 disease and CLN10 disease, respectively, which are two subtypes of neuronal ceroid lipofuscinosis (commonly known as Batten disease). The mechanisms regulating the intracellular trafficking of CLN5 and CTSD and their release from cells are not well understood. Here, we used the social amoeba Dictyostelium discoideum as a model system to examine the pathways and cellular components that regulate the intracellular trafficking and release of the D. discoideum homologs of human CLN5 (Cln5) and CTSD (CtsD). We show that both Cln5 and CtsD contain signal peptides for secretion that facilitate their release from cells. Like Cln5, extracellular CtsD is glycosylated. In addition, Cln5 release is regulated by the amount of extracellular CtsD. Autophagy induction promotes the release of Cln5, and to a lesser extent CtsD. Release of Cln5 requires the autophagy proteins Atg1, Atg5, and Atg9, as well as autophagosomal-lysosomal fusion. Atg1 and Atg5 are required for the release of CtsD. Together, these data support a model where Cln5 and CtsD are actively released from cells via their signal peptides for secretion and pathways linked to autophagy. The release of Cln5 and CtsD from cells also requires microfilaments and the D. discoideum homologs of human AP-3 complex mu subunit, the lysosomal-trafficking regulator LYST, mucopilin-1, and the Wiskott-Aldrich syndrome-associated protein WASH, which all regulate lysosomal exocytosis in this model organism. These findings suggest that lysosomal exocytosis also facilitates the release of Cln5 and CtsD from cells. In addition, we report the roles of ABC transporters, microtubules, osmotic stress, and the putative D. discoideum homologs of human sortilin and cation-independent mannose-6-phosphate receptor in regulating the intracellular/extracellular distribution of Cln5 and CtsD. In total, this study identifies the cellular mechanisms regulating the release of Cln5 and CtsD from D. discoideum cells and provides insight into how altered trafficking of CLN5 and CTSD causes disease in humans.

Endosomal vesicle fusion machinery is involved with the contractile vacuole in Dictyostelium discoideum.

Contractile vacuoles (CVs), enigmatic osmoregulatory organelles, share common characteristics, such as a requirement for RAB11 and high levels of V-ATPase. These commonalities suggest a conserved evolutionary origin for the CVs with implications for understanding of the last common ancestor of eukaryotes and eukaryotic diversification more broadly. A taxonomically broader sampling of CV-associated machinery is required to address this question further. We used a transcriptomics-based approach to identify CV-associated gene products in Dictyostelium discoideum. This approach was first validated by assessing a set of known CV-associated gene products, which were significantly upregulated following hypo-osmotic exposure. Moreover, endosomal and vacuolar gene products were enriched in the upregulated gene set. An upregulated SNARE protein (NPSNB) was predominantly plasma membrane localised and enriched in the vicinity of CVs, supporting the association with this organelle found in the transcriptomic analysis. We therefore confirm that transcriptomic approaches can identify known and novel players in CV function, in our case emphasizing the role of endosomal vesicle fusion machinery in the D. discoideum CV and facilitating future work to address questions regarding the deep evolution of eukaryotic organelles.

Cyclic di-AMP Signaling in Bacteria.

The second messenger molecule cyclic di-AMP (c-di-AMP) is formed by many bacteria and archaea. In many species that produce c-di-AMP, this second messenger is essential for viability on rich medium. Recent research has demonstrated that c-di-AMP binds to a large number of proteins and riboswitches, which are often involved in potassium and osmotic homeostasis. c-di-AMP becomes dispensable if the bacteria are cultivated on minimal media with low concentrations of osmotically active compounds. Thus, the essentiality of c-di-AMP does not result from an interaction with a single essential target but rather from the multilevel control of complex homeostatic processes. This review summarizes current knowledge on the homeostasis of c-di-AMP and its function(s) in the control of cellular processes.

Deciphering the function of the fifth class of Gα proteins: regulation of ionic homeostasis as unifying hypothesis.

Trimeric G proteins transduce signals from a superfamily of receptors and each G protein controls a wide range of cellular and systemic functions. Their highly conserved alpha subunits fall in five classes, four of which have been well investigated (Gs, Gi, G12, Gq). In contrast, the function of the fifth class, Gv is completely unknown, despite its broad occurrence and evolutionary ancient origin (older than metazoans). Here we show a dynamic presence of Gv mRNA in several organs during early development of zebrafish, including the hatching gland, the pronephros and several cartilage anlagen, employing in situ hybridisation. Next, we generated a Gv frameshift mutation in zebrafish and observed distinct phenotypes such as reduced oviposition, premature hatching and craniofacial abnormalities in bone and cartilage of larval zebrafish. These phenotypes could suggest a disturbance in ionic homeostasis as a common denominator. Indeed, we find reduced levels of calcium, magnesium and potassium in the larvae and changes in expression levels of the sodium potassium pump atp1a1a.5 and the sodium/calcium exchanger ncx1b in larvae and in the adult kidney, a major osmoregulatory organ. Additionally, expression of sodium chloride cotransporter slc12a3 and the anion exchanger slc26a4 is altered in complementary ways in adult kidney. It appears that Gv may modulate ionic homeostasis in zebrafish during development and in adults. Our results constitute the first insight into the function of the fifth class of G alpha proteins.

Managing fuels and fluids: Network integration of osmoregulatory and metabolic hormonal circuits in the polymodal control of homeostasis in insects.

Osmoregulation in insects is an essential process whereby changes in hemolymph osmotic pressure induce the release of diuretic or antidiuretic hormones to recruit individual osmoregulatory responses in a manner that optimizes overall homeostasis. However, the mechanisms by which different osmoregulatory circuits interact with other homeostatic networks to implement the correct homeostatic program remain largely unexplored. Surprisingly, recent advances in insect genetics have revealed several important metabolic functions are regulated by classic osmoregulatory pathways, suggesting that internal cues related to osmotic and metabolic perturbations are integrated by the same hormonal networks. Here, we review our current knowledge on the network mechanisms that underpin systemic osmoregulation and discuss the remarkable parallels between the hormonal networks that regulate body fluid balance and those involved in energy homeostasis to provide a framework for understanding the polymodal optimization of homeostasis in insects.

Discovery of prolactin-like in lamprey: Role in osmoregulation and new insight into the evolution of the growth hormone/prolactin family.

We used a representative of one of the oldest extant vertebrate lineages (jawless fish or agnathans) to investigate the early evolution and function of the growth hormone (GH)/prolactin (PRL) family. We identified a second member of the GH/PRL family in an agnathan, the sea lamprey (Petromyzon marinus). Structural, phylogenetic, and synteny analyses supported the identification of this hormone as prolactin-like (PRL-L), which has led to added insight into the evolution of the GH/PRL family. At least two ancestral genes were present in early vertebrates, which gave rise to distinct GH and PRL-L genes in lamprey. A series of gene duplications, gene losses, and chromosomal rearrangements account for the diversity of GH/PRL-family members in jawed vertebrates. Lamprey PRL-L is produced in the proximal pars distalis of the pituitary and is preferentially bound by the lamprey PRL receptor, whereas lamprey GH is preferentially bound by the lamprey GH receptor. Pituitary PRL-L messenger RNA (mRNA) levels were low in larvae, then increased significantly in mid-metamorphic transformers (stage 3); thereafter, levels subsided in final-stage transformers and metamorphosed juveniles. The abundance of PRL-L mRNA and immunoreactive protein increased in the pituitary of juveniles under hypoosmotic conditions, and treatment with PRL-L blocked seawater-associated inhibition of freshwater ion transporters. These findings clarify the origin and divergence of GH/PRL family genes in early vertebrates and reveal a function of PRL-L in osmoregulation of sea lamprey, comparable to a role of PRLs that is conserved in jawed vertebrates.

Genomic insights into the seawater adaptation in Cyprinidae.

BACKGROUND: Cyprinidae, the largest fish family, encompasses approximately 367 genera and 3006 species. While they exhibit remarkable adaptability to diverse aquatic environments, it is exceptionally rare to find them in seawater, with the Far Eastern daces being of few exceptions. Therefore, the Far Eastern daces serve as a valuable model for studying the genetic mechanisms underlying seawater adaptation in Cyprinidae. RESULTS: Here, we sequenced the chromosome-level genomes of two Far Eastern daces (Pseudaspius brandtii and P. hakonensis), the two known cyprinid fishes found in seawater, and performed comparative genomic analyses to investigate their genetic mechanism of seawater adaptation. Demographic history reconstruction of the two species reveals that their population dynamics are correlated with the glacial-interglacial cycles and sea level changes. Genomic analyses identified Pseudaspius-specific genetic innovations related to seawater adaptation, including positively selected genes, rapidly evolving genes, and conserved non-coding elements (CNEs). Functional assays of Pseudaspius-specific variants of the prolactin (prl) gene showed enhanced cell adaptation to greater osmolarity. Functional assays of Pseudaspius specific CNEs near atg7 and usp45 genes suggest that they exhibit higher promoter activity and significantly induced at high osmolarity. CONCLUSIONS: Our results reveal the genome-wide evidence for the evolutionary adaptation of cyprinid fishes to seawater, offering valuable insights into the molecular mechanisms supporting the survival of migratory fish in marine environments. These findings are significant as they contribute to our understanding of how cyprinid fishes navigate and thrive in diverse aquatic habitats, providing useful implications for the conservation and management of marine ecosystems.

Effects of CRISPR/Cas9 targeting of the myo-inositol biosynthesis pathway on hyper-osmotic tolerance of tilapia cells.

Myo-inositol is an important compatible osmolyte in vertebrates. This osmolyte is produced by the myo-inositol biosynthesis (MIB) pathway composed of myo-inositol phosphate synthase and inositol monophosphatase. These enzymes are among the highest upregulated proteins in tissues and cell cultures from teleost fish exposed to hyperosmotic conditions indicating high importance of this pathway for tolerating this type of stress. CRISPR/Cas9 gene editing of tilapia cells produced knockout lines of MIB enzymes and control genes. Metabolic activity decreased significantly for MIB KO lines in hyperosmotic media. Trends of faster growth of the MIB knockout lines in isosmotic media and faster decline of MIB knockout lines in hyperosmotic media were also observed. These results indicate a decline in metabolic fitness but only moderate effects on cell survival when tilapia cells with disrupted MIB genes are exposed to hyperosmolality. Therefore MIB genes are required for full osmotolerance of tilapia cells.

Adaptation and Survival of Marine-Associated Spiders (Araneae).

Aquatic environments are an unusual habitat for most arthropods. Nevertheless, many arthropod species that were once terrestrial dwelling have transitioned back to marine and freshwater environments, either as semiaquatic or, more rarely, as fully aquatic inhabitants. Transition to water from land is exceptional, and without respiratory modifications to allow for extended submergence and the associated hypoxic conditions, survival is limited. In this article, we review marine-associated species that have made this rare transition in a generally terrestrial group, spiders. We include several freshwater spider species for comparative purposes. Marine-associated spiders comprise less than 0.3% of spider species worldwide but are found in over 14% of all spider families. As we discuss, these spiders live in environments that, with tidal action, hydraulic forces, and saltwater, are more extreme than freshwater habitats, often requiring physiological and behavioral adaptations to survive. Spiders employ many methods to survive inundation from encroaching tides, such as air bubble respiration, airtight nests, hypoxic comas, and fleeing incoming tides. While airway protection is the primary survival strategy, further survival adaptations include saltwater-induced osmotic regulation, dietary composition, predator avoidance, reproduction, locomotory responses, and adaptation to extreme temperatures and hydrostatic pressures that challenge existence in marine environments.

Whitefly-Plant Interactions: An Integrated Molecular Perspective.

The rapid advances in available transcriptomic and genomic data and our understanding of the physiology and biochemistry of whitefly-plant interactions have allowed us to gain new and significant insights into the biology of whiteflies and their successful adaptation to host plants. In this review, we provide a comprehensive overview of the mechanisms that whiteflies have evolved to overcome the challenges of feeding on phloem sap. We also highlight the evolution and functions of gene families involved in host perception, evaluation, and manipulation; primary metabolism; and metabolite detoxification. We discuss the emerging themes in plant immunity to whiteflies, focusing on whitefly effectors and their sites of action in plant defense-signaling pathways. We conclude with a discussion of advances in the genetic manipulation of whiteflies and the potential that they hold for exploring the interactions between whiteflies and their host plants, as well as the development of novel strategies for the genetic control of whiteflies.

Mechanisms of Systemic Osmoregulation in Insects.

Water is essential to life. Terrestrial insects lose water by evaporation from the body surface and respiratory surfaces, as well as in the excretory products, posing a challenge made more acute by their high surface-to-volume ratio. These losses must be kept to a minimum and be offset by water gained from other sources. By contrast, insects such as the blood-sucking bug Rhodnius prolixus consume up to 10 times their body weight in a single blood meal, necessitating rapid expulsion of excess water and ions. How do insects manage their ion and water budgets? A century of study has revealed a great deal about the organ systems that insects use to maintain their ion and water balance and their regulation. Traditionally, a taxonomically wide range of species were studied, whereas more recent research has focused on model organisms to leverage the power of the molecular genetic approach. Key advances in new technologies have become available for a wider range of species in the past decade. We document how these approaches have already begun to inform our understanding of the diversity and conservation of insect systemic osmoregulation. We advocate that these technologies be combined with traditional approaches to study a broader range of nonmodel species to gain a comprehensive overview of the mechanism underpinning systemic osmoregulation in the most species-rich group of animals on earth, the insects.

Effects of pejus and pessimum zone salinity stress on gill proteome networks and energy homeostasis in Oreochromis mossambicus.

Salinity tolerance in fish involves a suite of physiological changes, but a cohesive theory leading to a mechanistic understanding at the organismal level is lacking. To examine the potential of adapting energy homeostasis theory in the context of salinity stress in teleost fish, Oreochromis mossambicus were acclimated to hypersalinity at multiple rates and durations to determine salinity ranges of tolerance and resistance. Over 3000 proteins were quantified simultaneously to analyze molecular phenotypes associated with hypersalinity. A species- and tissue-specific data-independent acquisition (DIA) assay library of MSMS spectra was created. Protein networks representing complex molecular phenotypes associated with salinity acclimation were generated. O. mossambicus has a wide "zone of resistance" from 75 g/kg salinity to 120 g/kg. Crossing into the zone of resistance resulted in marked phenotypic changes including blood osmolality over 400 mOsm/kg, reduced body condition, and cessation of feeding. Protein networks impacted by hypersalinity consist of electron transport chain (ETC) proteins and specific osmoregulatory proteins. Cytoskeletal, cell adhesion, and extracellular matrix proteins are enriched in networks that are sensitive to the critical salinity threshold. These network analyses identify specific proteome changes that are associated with distinct zones described by energy homeostasis theory and distinguish them from general hypersalinity-induced proteome changes.

NaX Channel Is a Physiological [Na+] Detector in Oxytocin- and Vasopressin-Releasing Magnocellular Neurosecretory Cells of the Rat Supraoptic Nucleus.

The Scn7A gene encodes NaX, an atypical noninactivating Na+ channel, whose expression in sensory circumventricular organs is essential to maintain homeostatic responses for body fluid balance. However, NaX has also been detected in homeostatic effector neurons, such as vasopressin (VP)-releasing magnocellular neurosecretory cells (MNCVP) that secrete VP (antidiuretic hormone) into the bloodstream in response to hypertonicity and hypernatremia. Yet, the physiological relevance of NaX expression in these effector cells remains unclear. Here, we show that rat MNCVP in males and females is depolarized and excited in proportion with isosmotic increases in [Na+]. These responses were caused by an inward current resulting from a cell-autonomous increase in Na+ conductance. The Na+-evoked current was unaffected by blockers of other Na+-permeable ion channels but was significantly reduced by shRNA-mediated knockdown of Scn7A expression. Furthermore, reducing the density of NaX channels selectively impaired the activation of MNCVP by systemic hypernatremia without affecting their responsiveness to hypertonicity in vivo These results identify NaX as a physiological Na+ sensor, whose expression in MNCVP contributes to the generation of homeostatic responses to hypernatremia.SIGNIFICANCE STATEMENT In this study, we provide the first direct evidence showing that the sodium-sensing channel encoded by the Scn7A gene (NaX) mediates cell-autonomous sodium detection by MNCs in the low millimolar range and that selectively reducing the expression of these channels in MNCs impairs their activation in response to a physiologically relevant sodium stimulus in vitro and in vivo These data reveal that NaX operates as a sodium sensor in these cells and that the endogenous sensory properties of osmoregulatory effector neurons contribute to their homeostatic activation in vivo.

TRPV4 and chloride channels mediate volume sensing in trabecular meshwork cells.

Aqueous humor drainage from the anterior eye determines intraocular pressure (IOP) under homeostatic and pathological conditions. Swelling of the trabecular meshwork (TM) alters its flow resistance but the mechanisms that sense and transduce osmotic gradients remain poorly understood. We investigated TM osmotransduction and its role in calcium and chloride homeostasis using molecular analyses, optical imaging, and electrophysiology. Anisosmotic conditions elicited proportional changes in TM cell volume, with swelling, but not shrinking, evoking elevations in intracellular calcium concentration [Ca2+]TM. Hypotonicity-evoked calcium signals were sensitive to HC067047, a selective blocker of TRPV4 channels, whereas the agonist GSK1016790A promoted swelling under isotonic conditions. TRPV4 inhibition partially suppressed hypotonicity-induced volume increases and reduced the magnitude of the swelling-induced membrane current, with a substantial fraction of the swelling-evoked current abrogated by Cl- channel antagonists 4,4'-diisothiocyanato-2,2'-stilbenedisulfonic acid (DIDS) and niflumic acid. The transcriptome of volume-sensing chloride channel candidates in primary human was dominated by ANO6 transcripts, with moderate expression of ANO3, ANO7, and ANO10 transcripts and low expression of LTTRC genes that encode constituents of the volume-activated anion channel. Imposition of 190 mosM but not 285 mosM hypotonic gradients increased conventional outflow in mouse eyes. TRPV4-mediated cation influx thus works with Cl- efflux to sense and respond to osmotic stress, potentially contributing to pathological swelling, calcium overload, and intracellular signaling that could exacerbate functional disturbances in inflammatory disease and glaucoma.NEW & NOTEWORTHY Intraocular pressure is dynamically regulated by the flow of aqueous humor through paracellular passages within the trabecular meshwork (TM). This study shows hypotonic gradients that expand the TM cell volume and reduce the outflow facility in mouse eyes. The swelling-induced current consists of TRPV4 and chloride components, with TRPV4 as a driver of swelling-induced calcium signaling. TRPV4 inhibition reduced swelling, suggesting a novel treatment for trabeculitis and glaucoma.

Staying Hydrated in Seawater.

Reduction of intestinal lumen osmotic pressure by the formation of Ca(Mg)CO3, "ichthyocarbonate," is essential for osmoregulation by the only vertebrate group, ray-finned fishes, widely capable of hydrating by ingesting seawater. Ichthyocarbonate formation and excretion are under elaborate physiological control and play an important, yet still poorly defined, role in the oceanic carbon cycle.

Multivariate investigation of Moringa oleifera morpho-physiological and biochemical traits under various water regimes.

BACKGROUND: The climatic changes crossing the world menace the green life through limitation of water availability. The goal of this study was to determine whether Moringa oleifera Lam. trees cultivated under Tunisian arid climate, retain their tolerance ability to tolerate accentuated environmental stress factors such as drought and salinity. For this reason, the seeds of M. oleifera tree planted in Bouhedma Park (Tunisian arid area), were collected, germinated, and grown in the research area at the National Institute of Research in Rural Engineering, Waters and Forests (INRGREF) of Tunis (Tunisia). The three years aged trees were exposed to four water-holding capacities (25, 50, 75, and 100%) for 60 days to realise this work. RESULTS: Growth change was traduced by the reduction of several biometric parameters and fluorescence (Fv/Fm) under severe water restriction (25 and 50%). Whereas roots presented miraculous development in length face to the decrease of water availability (25 and 50%) in their rhizospheres. The sensitivity to drought-induced membrane damage (Malondialdehyde (MDA) content) and reactive oxygen species (ROS) liberation (hydrogen peroxide (H2O2) content) was highly correlated with ROS antiradical scavenging (ferric reducing antioxidant power (FRAP) and (2, 2'-diphenyl-1-picrylhydrazyle (DPPH)), phenolic components and osmolytes accumulation. The drought stress tolerance of M. oleifera trees was associated with a dramatic stimulation of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), ascorbate peroxidase (APX), and glutathione peroxidase (GPX) activities. CONCLUSION: Based on the several strategies adopted, integrated M. oleifera can grow under drought stress as accentuated adverse environmental condition imposed by climate change.

Biallelic loss-of-function variants of SLC12A9 cause lysosome dysfunction and a syndromic neurodevelopmental disorder.

PURPOSE: Pathogenic variants of FIG4 generate enlarged lysosomes and neurological and developmental disorders. To identify additional genes regulating lysosomal volume, we carried out a genome-wide activation screen to detect suppression of enlarged lysosomes in FIG4-/- cells. METHODS: The CRISPR-a gene activation screen utilized sgRNAs from the promoters of protein-coding genes. Fluorescence-activated cell sorting separated cells with correction of the enlarged lysosomes from uncorrected cells. Patient variants of SLC12A9 were identified by exome or genome sequencing and studied by segregation analysis and clinical characterization. RESULTS: Overexpression of SLC12A9, a solute co-transporter, corrected lysosomal swelling in FIG4-/- cells. SLC12A9 (NP_064631.2) colocalized with LAMP2 at the lysosome membrane. Biallelic variants of SLC12A9 were identified in 3 unrelated probands with neurodevelopmental disorders. Common features included intellectual disability, skeletal and brain structural abnormalities, congenital heart defects, and hypopigmented hair. Patient 1 was homozygous for nonsense variant p.(Arg615∗), patient 2 was compound heterozygous for p.(Ser109Lysfs∗20) and a large deletion, and proband 3 was compound heterozygous for p.(Glu290Glyfs∗36) and p.(Asn552Lys). Fibroblasts from proband 1 contained enlarged lysosomes that were corrected by wild-type SLC12A9 cDNA. Patient variant p.(Asn552Lys) failed to correct the lysosomal defect. CONCLUSION: Impaired function of SLC12A9 results in enlarged lysosomes and a recessive disorder with a recognizable neurodevelopmental phenotype.

Molecular basis of the unique osmoregulatory strategy in the inshore hagfish, Eptatretus burgeri.

Hagfishes are characterized by omo- and iono-conforming nature similar to marine invertebrates. Conventionally, hagfishes had been recognized as the most primitive living vertebrate that retains plesiomorphic features. However, some of the "ancestral" features of hagfishes, such as rudimentary eyes and the lack of vertebrae, have been proven to be deceptive. Similarly, by the principle of maximum parsimony, the unique body fluid regulatory strategy of hagfishes seems to be apomorphic, since the lamprey, another cyclostome, adopts osmo- and iono-regulatory mechanisms as in jawed vertebrates. Although hagfishes are unequivocally important in discussing the origin and evolution of the vertebrate osmoregulatory system, the molecular basis for the body fluid homeostasis in hagfishes has been poorly understood. In the present study, we explored this matter in the inshore hagfish, Eptatretus burgeri, by analyzing the transcriptomes obtained from the gill, kidney, and muscle of the animals acclimated to distinct environmental salinities. Together with the measurement of parameters in the muscular fluid compartment, our data indicate that the hagfish possesses an ability to conduct free amino acid (FAA)-based osmoregulation at a cellular level, which is in coordination with the renal and branchial FAA absorption. We also revealed that the hagfish does possess the orthologs of the known osmoregulatory genes and that the transepithelial movement of inorganic ions in the hagfish gill and kidney is more complex than previously thought. These observations pose a challenge to the conventional view that the physiological features of hagfishes have been inherited from the last common ancestor of the extant vertebrates.

Responses of Microorganisms to Osmotic Stress.

The cytoplasm of bacterial cells is a highly crowded cellular compartment that possesses considerable osmotic potential. As a result, and owing to the semipermeable nature of the cytoplasmic membrane and the semielastic properties of the cell wall, osmotically driven water influx will generate turgor, a hydrostatic pressure considered critical for growth and viability. Both increases and decreases in the external osmolarity inevitably trigger water fluxes across the cytoplasmic membrane, thus impinging on the degree of cellular hydration, molecular crowding, magnitude of turgor, and cellular integrity. Here, we assess mechanisms that permit the perception of osmotic stress by bacterial cells and provide an overview of the systems that allow them to genetically and physiologically cope with this ubiquitous environmental cue. We highlight recent developments implicating the secondary messenger c-di-AMP in cellular adjustment to osmotic stress and the role of osmotic forces in the life of bacteria-assembled in biofilms.

Environmental salinity modulates olfactory sensitivity in the euryhaline European seabass, Dicentrarchus labrax, acclimated to seawater and brackish water.

The olfactory epithelium of fish is - of necessity - in intimate contact with the surrounding water. In euryhaline fish, movement from seawater to freshwater (and vice versa) exposes the epithelium to massive changes in salinity and ionic concentrations. How does the olfactory system function in the face of such changes? The current study compared olfactory sensitivity in seawater- (35‰) and brackish water-adapted seabass (5‰) using extracellular multi-unit recording from the olfactory nerve. Seawater-adapted bass had higher olfactory sensitivity to amino acid odorants when delivered in seawater than in freshwater. Conversely, brackish water-adapted bass had largely similar sensitivities to the same odorants when delivered in seawater or freshwater, although sensitivity was still slightly higher in seawater. The olfactory system of seawater-adapted bass was sensitive to decreases in external [Ca2+], whereas brackish water-adapted bass responded to increases in [Ca2+]; both seawater- and brackish water-adapted bass responded to increases in external [Na+] but the sensitivity was markedly higher in brackish water-adapted bass. In seawater-adapted bass, olfactory sensitivity to l-alanine depended on external Ca2+ ions, but not Na+; brackish water-adapted bass did respond to l-alanine in the absence of Ca2+, albeit with lower sensitivity, whereas sensitivity was unaffected by removal of Na+ ions. A possible adaptation of the olfactory epithelium was the higher number of mucous cells in brackish water-adapted bass. The olfactory system of seabass is able to adapt to low salinities, but this is not immediate; further studies are needed to identify the processes involved.