The Cargo Receptor NDP52 Initiates Selective Autophagy by Recruiting the ULK Complex to Cytosol-Invading Bacteria.

Xenophagy, a selective autophagy pathway that protects the cytosol against bacterial invasion, relies on cargo receptors that juxtapose bacteria and phagophore membranes. Whether phagophores are recruited from a constitutive pool or are generated de novo at prospective cargo remains unknown. Phagophore formation in situ would require recruitment of the upstream autophagy machinery to prospective cargo. Here, we show that, essential for anti-bacterial autophagy, the cargo receptor NDP52 forms a trimeric complex with FIP200 and SINTBAD/NAP1, which are subunits of the autophagy-initiating ULK and the TBK1 kinase complex, respectively. FIP200 and SINTBAD/NAP1 are each recruited independently to bacteria via NDP52, as revealed by selective point mutations in their respective binding sites, but only in their combined presence does xenophagy proceed. Such recruitment of the upstream autophagy machinery by NDP52 reveals how detection of cargo-associated "eat me" signals, induction of autophagy, and juxtaposition of cargo and phagophores are integrated in higher eukaryotes.

Galectin-8 inhibition and functions in immune response and tumor biology.

Galectins are among organisms' most abundantly expressed lectins (carbohydrate-binding proteins) that specifically bind ß-galactosides. They act not only outside the cell, where they bind to extracellular matrix glycans, but also inside the cell, where they have a significant impact on signaling pathways. Galectin-8 is a galectin family protein encoded by the LGALS8 gene. Its role is evident in both T- and B-cell immunity and in the innate immune response, where it acts directly on dendritic cells and induces some pro-inflammatory cytokines. Galectin-8 also plays an important role in the defense against bacterial and viral infections. It is known to promote antibacterial autophagy by recognizing and binding glycans present on the vacuolar membrane, thus acting as a danger receptor. The most important role of galectin-8 is the regulation of cancer growth, metastasis, tumor progression, and tumor cell survival. Importantly, the expression of galectins is typically higher in tumor tissues than in noncancerous tissues. In this review article, we focus on galectin-8 and its function in immune response, microbial infections, and cancer. Given all of these functions of galectin-8, we emphasize the importance of developing new and selective galectin-8 inhibitors and report the current status of their development.

Lipid raft interacting galectin 8 regulates primary ciliogenesis.

Primary cilium is a specialized sensory organelle that transmits environmental information into cells. Its length is tightly controlled by various mechanisms such as the frequency or the cargo size of the intraflagellar transport trains which deliver the building materials such as tubulin subunits essential for the growing cilia. Here, we show the sialoglycan interacting galectin 8 regulates the process of primary ciliogenesis. As the epithelia become polarized, there are more galectin 8 being apically secreted and these extracellular galectin 8 molecules apparently bind to a lipid raft enriched domain at the base of the primary cilia through interacting with lipid raft components, such as GD3 ganglioside and scaffold protein caveolin 1. Furthermore, the binding of galectin 8 at this critical region triggers rapid growth of primary cilia by perturbing the barrier function of the transition zone (TZ). Our study also demonstrates the functionality of this barrier depends on intact organization of lipid rafts at the cilia as genetically knockout of Cav1 and pharmacologically inhibition of lipid raft both phenocopy the effect of apical addition of recombinant galectin 8; that is, rapid elongation of primary cilia and redistribution of cilia proteins from TZ to the growing axoneme. Indeed, as cilia elongated, endogenous galectin 8, caveolin 1, and TZ component, TMEM231, also transited from the TZ to the growing axoneme. We also noted that the interaction between caveolin 1 and TMEM231 could be perturbed by exogenous galectin 8. Taken together, we proposed that galectin 8 promoted primary cilia elongation through impeding the barrier function of the TZ by interfering with the interaction between caveolin 1 and TMEM231.

Unveiling the molecular characteristics and antibacterial activity of tandem-repeat-type Galectin-8 in large yellow croaker (Larimichthys crocea).

Galectin-8 (Gal-8) is a versatile carbohydrate-binding protein with pivotal roles in immune regulation and cellular processes. This study introduces a novel galectin-8 protein, LcGal-8, from the large yellow croaker (Larimichthys crocea), showcasing typical characteristics of tandem-repeat-type galectins, including the absence of a signal peptide or transmembrane region and the presence of conserved sugar-binding motifs. Phylogenetic analysis reveals its conservation among fish species. Expression profiling indicates widespread distribution in immune tissues, particularly the spleen, implicating involvement in immune processes. The subcellular localization analysis reveals that LcGal-8 is present in both the cytoplasm and nucleus. Upon bacterial challenge, LcGal-8 is up-regulated in immune tissues, suggesting a role in host defense. Functional assays demonstrate that LcGal-8 can agglutinate gram-negative bacteria. The recombinant LcGal-8 protein agglutinates red blood cells from the large yellow croaker independently of Ca2⁺, however, this activity is inhibited by lipopolysaccharide (LPS) at 2.5 µg/mL. Fluorescence detection kits and scanning electron microscopy (SEM) confirm the agglutination and bactericidal effects of LcGal-8 against various gram-negative bacteria, including Vibrio harveyi, Aeromondaceae hydrophila, Aeromondaceae veronii, Pseudomonas plecoglossicida, Edwardsiella tarda. These findings contribute valuable insights into the genetic basis of disease resistance in the large yellow croaker and could support molecular breeding strategies to enhance disease resistance.

Galectin-8 Contributes to Human Trophoblast Cell Invasion.

Galectins are a class of lectins that are extensively expressed in all organisms. Galectins are involved in a range of functions, including early development, tissue regeneration, cancer and inflammation. It has been shown that galectin-8 is expressed in the villous and extravillous trophoblast (EVT) cells of the human placenta; however, its physiological role in pregnancy establishment has not been elucidated. Taking these factors into account, we investigated the functional role of galectin-8 in HTR-8/SVneo cells-a human EVT cell line-and human primary cytotrophoblast cells isolated from a first-trimester placenta. We analyzed the effects of recombinant human galectin-8 (rh galectin-8) on the adhesion, migration and invasion of HTR-8/SVneo cells. We used qPCR, cell-based ELISA (cELISA) and gelatin zymography to study the effects of galectin-8 on mediators of these processes, such as integrin subunits alpha-1 and beta-1 and matrix metalloproteinases (MMPs)-2 and -9, on the mRNA and protein levels. Further, we studied the effects of galectin-8 on primary cytotrophoblast cells' invasion. Galectin-8 stimulated the adhesion, migration and invasion of HTR-8/SVneo cells, as well as the invasion of primary cytotrophoblasts. In addition, the MMP-2 and -9 levels were increased, while the expression of integrins alpha-1 and beta-1 was not affected. Galectin-8 has the ability to positively affect EVTs' invasion, so it can be considered a significant factor in the trophoblast cell invasion process.

Galectin-8 involves in arthritic condylar bone loss via podoplanin/AKT/ERK axis-mediated inflammatory lymphangiogenesis.

OBJECTIVE: The lymphatic system plays a crucial role in the maintenance of tissue fluid homeostasis and the immunological response to inflammation. Galectin-8 (Gal-8) regulates pathological lymphangiogenesis but the effects of which on inflammation-related condylar bone loss in temporomandibular joint (TMJ) have not been well studied. DESIGN: We used TNFα-transgenic (TNFTG) mice and their wildtype (WT) littermates to compare their inflammatory phenotype in TMJs. Next, lymphatic endothelial cells (LECs) were used to examine the effects of which on osteoclast formation, pro-inflammatory factor expression, and inflammatory lymphangiogenesis with or without thiodigalactoside (TDG, a Gal-8 inhibitor) treatment. At last, two murine models (TNFTG arthritic model and forced mouth opening model) were used to explore TDG as a potential drug for the treatment of inflammation-related condylar bone loss. RESULTS: In comparison to WT mice, lymphatic areas of lymphatic vessel endothelial receptor 1 (LYVE1)+/podoplanin (PDPN)+ and Gal-8+/PDPN+, TRAP-positive osteoclast number, and condylar bone loss are increased in TNFTG mice. Inhibition of Gal-8 in LECs by TDG, reduces TNFα-induced osteoclast formation, pro-inflammatory factor expression, and inflammatory lymphangiogenesis. In addition, Gal-8 promotes TNFα-activated AKT/ERK/NF-κB pathways by binding to PDPN. Finally, the administration of TDG attenuates inflammatory lymphangiogenesis, inhibits osteoclast activity, and reduces condylar bone loss in TNFTG arthritic mice and forced mouth opening mice. CONCLUSIONS: Our findings reveal the important role of Gal-8-promoted pathological lymphangiogenesis in inflammation-related condylar bone loss.

Galectin-8, cytokines, and the storm.

Galectin-8 (Gal-8) belongs to a family of animal lectins that modulate cell adhesion, cell proliferation, apoptosis, and immune responses. Recent studies have shown that mammalian Gal-8 induces in an autocrine and paracrine manner, the expression and secretion of cytokines and chemokines such as RANKL, IL-6, IL-1ß, SDF-1, and MCP-1. This involves Gal-8 binding to receptor complexes that include MRC2/uPAR/LRP1, integrins, and CD44. Receptors ligation triggers FAK, ERK, Akt, and the JNK signaling pathways, leading to induction of NF-κB that promotes cytokine expression. Indeed, immune-competent Gal-8 knockout (KO) mice express systemic lower levels of cytokines and chemokines while the opposite is true for Gal-8 transgenic animals. Cytokine and chemokine secretion, induced by Gal-8, promotes the migration of cancer cells toward cells expressing this lectin. Accordingly, Gal-8 KO mice experience reduced tumor size and smaller and fewer metastatic lesions when injected with cancer cells. These observations suggest the existence of a 'vicious cycle' whereby Gal-8 expression and secretion promotes the secretion of cytokines and chemokines that further promote Gal-8 expression. This 'vicious cycle' could enhance the development of a 'cytokine storm' which is a key contributor to the poor prognosis of COVID-19 patients.

Molecular characterization and expression profiling of tandem-repeat galectin-8 from red-spotted grouper (Epinephelus akaara): Potential antibacterial, antiviral, and wound healing activities.

Galectin-8 is a typical ß-galactoside binding lectin, which primarily functions as a pattern recognition receptor and/or danger receptor that is engaged in pathogen recognition by the host innate immune system. Although several fish galectins have been identified, the role of galectin-8 in teleost immunity is still not fully understood. In this study, molecular, transcriptional, and immune-related functions of galectin-8 (EaGal8) from red-spotted grouper (Epinephelus akaara) were analyzed. The open reading frame of EaGal8 comprised 960 bp encoding 319 amino acids of a ∼35 kDa protein, composed of the N- and C-terminal carbohydrate recognition domains joined by a short hinge peptide. Phylogenetic analysis revealed that EaGal8 was closely related to the Epinephelus lanceolatus galectin-8-like protein. Although EaGal8 showed ubiquitous tissue expression, the highest expression level was observed in the blood. Immunostimulants, including lipopolysaccharide, poly(I:C), and nervous necrosis virus, significantly upregulated the EaGal8 transcription level in a time-dependent manner (p < 0.05). Furthermore, recombinant EaGal8 (rEaGal8) showed a binding affinity toward seven different carbohydrates in a concentration-dependent manner. In addition, rEaGal8 caused strong agglutination of fish red blood cells and several gram-positive and gram-negative bacteria, including Streptococcus iniae, Streptococcus parauberis, Lactococcus garvieae, Escherichia coli, Edwardsiella tarda, Vibrio alginolyticus, Vibrio parahaemolyticus, and Pseudomonas aeruginosa. For the first time in teleosts, we report the wound healing ability of galectin-8 in this study. At low concentrations, rEaGal8 showed potential wound healing responses in FHM cells, in vitro. Thus, this study reinforces the role of EaGal8 in innate immune responses against bacterial and viral infections and wound healing in red-spotted grouper.

Galectin-8 and -9 as prognostic factors for cervical cancer.

PURPOSE: Galectins are carbohydrate-binding proteins with multiple effects on cell biology. Research shows that they play an important role in tumor development and progression. Therefore, in this study, the presence of Galectin-8 and -9 (Gal), both already known as prognostic factors in other tumor entities, were investigated in cervical cancer. Our aim was to examine the association of Gal-8 and -9 expression with histopathological markers and survival of the patients. METHODS: Gal-8 and -9 expression was investigated in 250 cervical cancer samples by immunohistochemistry. The staining was evaluated using the immunoreactive score (IRS). The results were correlated to clinical and pathological data. The correlation of Gal-8 and -9 expression with overall and relapse-free survival was analyzed. RESULTS: Expression of Gal-8 was associated with negative N-status and lower FIGO status. Detection of Gal-9 was connected to negative N-status and lower grading regarding all specimens. A correlation of Gal-9 with lower FIGO status was detected for squamous cell carcinoma (SCC) only. Expression of Gal-8 was associated with relapse-free survival of SCC patients in a positive manner. Gal-9 expression was associated with better overall survival. CONCLUSION: Our results suggest that expression of both galectins is inversely associated with tumor stage and progression. Gal-8 expression is associated with relapse-free survival of patients with SCC, while presence of Gal-9 in cervical cancer is associated with a better prognosis in regard of overall survival.

BST2, a Novel Inhibitory Receptor, Is Involved in NK Cell Cytotoxicity through Its Cytoplasmic Tail Domain.

Bone Marrow Stromal Cell Antigen 2 (BST2) is a type II transmembrane protein expressed on various cell types that tethers the release of viruses. Natural killer (NK) cells express low levels of BST2 under normal conditions but exhibit increased expression of BST2 upon activation. In this study, we show for the first time that murine BST2 can control the cytotoxicity of NK cells. The cytoplasmic tail of murine BST2 contains an immunoreceptor tyrosine-based inhibitory motif (ITIM). The absence of BST2 on NK cells can enhance their cytotoxicity against tumor cells compared to wild type NK cells. NK cells isolated from NZW mice, which express ITIM-deficient BST2, also showed higher cytotoxicity than wild type NK cells. In addition, we found that galectin-8 and galectin-9 were ligands of BST2, since blocking galectin-8 or -9 with monoclonal antibodies enhanced the cytotoxicity of NK cells. These results suggested that BST2 might be a novel NK cell inhibitory receptor as it was involved in regulating NK cell cytotoxicity through its interaction with galectins.

Molecular characterization, expression analysis and immune effect of Galectin-8 from Japanese flounder (Paralichthys olivaceus).

Galectin-8 gene belongs to the agglutinin family, which can specifically recognize ß-galactoside bonds and play essential roles in many biological processes. In this study, we researched the sequence characteristics and immune-related function of Galectin-8 gene in Japanese flounder Paralichthys olivaceus, named PoGalectin-8. The results showed that the open reading frame of PoGalectin-8 was 891 bp, which encoding a protein with 296 amino acid residues and containing typical HXNPR and WGXEE motifs in the N-terminal and C-terminal CRD domains. Sequence alignment showed that PoGalectin-8 was conserved in different aquatic animals and exhibited the highest similarity (95.27%) with Seriola dumerili. PoGalectin-8 expressed in all detected tissues and exhibited the highest expression level in spleen, followed by skin and kidney. After infected by Edwardsiella tarda, the expression of PoGalectin-8 was down-regulated in the spleen and skin tissues of P. olivaceus. Further to study its immune-related functions, the recombinant PoGalectin-8 (rPoGalectin-8) was expressed and purified. The rPoGalectin-8 can specifically bind to lipopolysaccharide and peptidoglycan, the main components of cell walls from Gram-negative and Gram-positive bacteria. Bacteria binding and the microbial agglutinating experiments showed that the rPoGalectin-8 could bind and agglutinate all examined Gram-positive and Gram-negative bacteria. This study implied that PoGalectin-8, as a pattern recognition receptor, may play important roles during immune responses against bacterial infection, which laid a foundation for further functional identification of Galectin-8 in aquatic animal immunity.

Full-length galectin-8 and separate carbohydrate recognition domains: the whole is greater than the sum of its parts?

Galectin-8 (Gal-8) is a tandem-repeat type galectin with affinity for ß-galactosides, bearing two carbohydrate recognition domains (CRD) connected by a linker peptide. The N- and C-terminal domains (Gal-8N and Gal-8C) share 35% homology, and their glycan ligand specificity is notably dissimilar: while Gal-8N shows strong affinity for α(2-3)-sialylated oligosaccharides, Gal-8C has higher affinity for non-sialylated oligosaccharides, including poly-N-acetyllactosamine and/ or A and B blood group structures. Particularly relevant for understanding the biological role of this lectin, full-length Gal-8 can bind cell surface glycoconjugates with broader affinity than the isolated Gal-8N and Gal-8C domains, a trait also described for other tandem-repeat galectins. Herein, we aim to discuss the potential use of separate CRDs in modelling tandem-repeat galectin-8 and its biological functions. For this purpose, we will cover several aspects of the structure-function relationship of this protein including crystallographic structures, glycan specificity, cell function and biological roles, with the ultimate goal of understanding the potential role of each CRD in predicting full-length Gal-8 involvement in relevant biological processes.

Expression and antibacterial analysis of galectin-8 and -9 genes in mandarin fish, Siniperca chuatsi.

Galectin-8 and galectin-9 belong to tandem repeat-type galectins, and in the present study, these two genes were cloned in mandarin fish Siniperca chuatsi. The open reading frame (ORF) of the mandarin fish galectin-8 and galectin-9 contains 942, and 1008 bp, encoding 313 and 335 amino acids, respectively. As a conserved feature, an N-terminal carbohydrate recognition domain (CRD), and a C-terminal CRD were observed in each of the two galectins in mandarin fish. In healthy fish, galectin-8 and -9 were constitutively expressed in all organs/tissues examined, and their expression can be induced following the stimulation of LPS and poly(I:C). It is obvious that galectin-8 had a higher increase at mRNA level following the stimulation of poly(I:C). It is further demonstrated that mandarin fish galectin-8 inhibited the growth of Flavobacterium columnare and Streptococcus agalactiae, and in addition to the two species of bacteria, galectin-9 inhibited also the growth of Edwardsiella piscicida, which provides the basis for further understanding their antibacterial role in immune response of mandarin fish.

A Fish Galectin-8 Possesses Direct Bactericidal Activity.

Galectins are a family of animal lectins with high affinity for ß-galactosides. Galectins are able to bind to bacteria, and a few mammalian galectins are known to kill the bound bacteria. In fish, no galectins with direct bactericidal effect have been reported. In the present study, we identified and characterized a tandem repeat galectin-8 from tongue sole Cynoglossus semilaevis (designated CsGal-8). CsGal-8 possesses conserved carbohydrate recognition domains (CRDs), as well as the conserved HXNPR and WGXEE motifs that are critical for carbohydrate binding. CsGal-8 was constitutively expressed in nine tissues of tongue sole and up-regulated in kidney, spleen, and blood by bacterial challenge. When expressed in HeLa cells, CsGal-8 protein was detected both in the cytoplasm and in the micro-vesicles secreted from the cells. Recombinant CsGal-8 (rCsGal-8) bound to lactose and other carbohydrates in a dose dependent manner. rCsGal-8 bound to a wide range of gram-positive and gram-negative bacteria and was co-localized with the bound bacteria in animal cells. Lactose, fructose, galactose, and trehalose effectively blocked the interactions between rCsGal-8 and different bacteria. Furthermore, rCsGal-8 exerted potent bactericidal activity against some gram-negative bacterial pathogens by directly damaging the membrane and structure of the pathogens. Taken together, these results indicate that CsGal-8 likely plays an important role in the immune defense against some bacterial pathogens by direct bacterial interaction and killing.

Galectin-8-mediated selective autophagy protects against seeded tau aggregation.

Assembled tau can transfer between cells and seed the aggregation of soluble tau. This process is thought to underlie the amplification and propagation of tau inclusions throughout the brain in neurodegenerative diseases, including Alzheimer's disease. An understanding of the mechanisms involved may provide strategies for limiting assembled tau propagation. Here, we sought to determine how assembled tau seeds gain access to the cytosol and whether this access triggers cellular defenses. We show that tau assemblies enter cells through clathrin-independent endocytosis and escape from damaged endomembranes into the cytosol, where they seed the aggregation of soluble tau. We also found that the danger receptor galectin-8 detects damaged endomembranes and activates autophagy through recruitment of the cargo receptor nuclear dot protein 52 (NDP52). Inhibition of galectin-8- and NDP52-dependent autophagy increased seeded tau aggregation, indicating that autophagy triggered by damaged endomembranes during the entry of assembled tau seeds protects against tau aggregation, in a manner similar to cellular defenses against cytosol-dwelling microorganisms. A second autophagy cargo receptor, p62, then targeted seeded tau aggregates. Our results reveal that by monitoring endomembrane integrity, cells reduce entry of tau seeds into the cytosol and thereby prevent seeded aggregation. The mechanisms described here may help inform the development of therapies aimed at inhibiting the propagation of protein assemblies in neurodegenerative diseases.

Intracellular galectins control cellular responses commensurate with cell surface carbohydrate composition.

Galectins are ß-galactoside-binding animal lectins primarily found in the cytosol, while their carbohydrate ligands are mainly distributed in the extracellular space. Cytosolic galectins are anticipated to accumulate on damaged endocytic vesicles through binding to glycans initially displayed on the cell surface and subsequently located in the lumen of the vesicles, and this can be followed by cellular responses. To facilitate elucidation of the mechanism underlying this process, we adopted a model system involving induction of endocytic vesicle damage with light that targets the endocytosed amphiphilic photosensitizer disulfonated aluminum phthalocyanine. We demonstrate that the levels of galectins around damaged endosomes are dependent on the composition of carbohydrates recognized by the proteins. By super resolution imaging, galectin-3 and galectin-8 aggregates were found to be distributed in distinct microcompartments. Importantly, galectin accumulation is significantly affected when cell surface glycans are altered. Furthermore, accumulated galectins can direct autophagy adaptor proteins toward damaged endocytic vesicles, which are also significantly affected following alteration of cell surface glycans. We conclude that cytosolic galectins control cellular responses reflect dynamic modifications of cell surface glycans.

Kojic acid inhibits senescence of human corneal endothelial cells via NF-κB and p21 signaling pathways.

Fuchs endothelial dystrophy (FED) and late cornea allograft failure of cornea transplantation are associated with human corneal endothelial cells (HCEC) senescence. Kojic acid has various functions, however, its anti-senescence effect has never been identified. In this study, we investigated the anti-senescence effect of kojic acid on HCEC. Cell viability, migration ability and senescence were evaluated by MTT assay, migration assay, and senescence-associated beta-galactosidase (SA-ß-Gal) staining, respectively. Senescence-related protein expression was analyzed by western blotting and immunofluorescence assay. Angiogenesis of human umbilical vein endothelial cells (HUVEC) was examined by tube formation assay and spheroid sprouting assay. The results showed that kojic acid could inhibit HCEC senescence, characterized by enhancing migration, decreasing the levels of SA-ß-Gal staining, galectin 8, laminin α1, laminin α2, laminin γ1 and p21, and increasing that of p-NF-κB of senescent HCEC. The p-NF-κB inhibitor could reverse the anti-senescent effect of kojic acid, and p21 siRNA showed similar anti-senescence effect with kojic acid. In addition, kojic acid could alleviate HUVEC tube formation induced by senescent HCEC, which could be reversed by p-NF-κB inhibitor. The p21 siRNA could alleviate HUVEC spheroid sprouting induced by senescent HCEC. These results indicated that kojic acid might inhibit HCEC senescence and following resulted angiogenesis via NF-κB and p21 signaling pathways, possibly through downregulation of galectin 8 and laminins. Therefore, kojic acid is a promising drug for HCEC senescence-related diseases such as FED and late cornea allograft failure.

Molecular, transcriptional and functional delineation of Galectin-8 from black rockfish (Sebastes schlegelii) and its potential immunological role.

Galectins are ß-galactoside-binding lectins, which are involved in pattern recognition, cell adhesion, and stimulation of the host innate immune responses against microbial pathogens. In spite of several functional studies on different galectins isolated from vertebrates and invertebrates, this is the first report to present functional studies for galectin-8 from the marine teleost tissues. In the present study, we characterized galectin-8 homolog from black rockfish (Sebastes schlegelii), in molecular and functional aspects. Rockfish galectin-8 (SsGal8) was found to consist of a 969 bp long open reading frame (ORF), encoding a protein of 322 amino acids and the predicted molecular weight was 35.82 kDa. In silico analysis of SsGal8 revealed the presence of two carbohydrate binding domains (CRDs), at both N and C-termini and a linker peptide of 40 amino acids, in between the two domains. As expected, the phylogenetic tree categorized SsGal8 as a tandem-repeat galectin, and ultimately positioned it in the sub-clade of fish galectin-8. rSsGal8 was able to strongly agglutinate fish erythrocytes and the inhibition of agglutination was successfully exhibited by lactose and d-galactose. Bacterial agglutination assay resulted in agglutination of both Gram (+) and Gram (-) bacteria, including Escherichia coli, Vibrio harveyi, Vibrio parahaemolyticus, Streptococcus parauberis, Lactococcus garvieae, Streptococcus iniae and Vibrio tapetis. The tissue distribution analysis based on qPCR assays, revealed a ubiquitous tissue expression of SsGal8 for the examined rockfish tissues, with the most pronounced expression in blood, followed by brain, intestine, head kidney and kidney. Furthermore, the mRNA transcription level of SsGal8 was significantly up-regulated in spleen, liver and head kidney, upon immune challenges with Streptococcus iniae, LPS and poly I:C, in a time dependent manner. Taken together, these findings strongly suggest the contribution of SsGal8 in regulating innate immune responses to protect the rockfish from bacterial infections.

Overall Survival of Ovarian Cancer Patients Is Determined by Expression of Galectins-8 and -9.

The evaluation of new prognostic factors that can be targeted in ovarian cancer diagnosis and therapy is of the utmost importance. Galectins are a family of carbohydrate binding proteins with various implications in cancer biology. In this study, the presence of galectin (Gal)-8 and -9 was investigated in 156 ovarian cancer samples using immunohistochemistry (IHC). Staining was evaluated using semi-quantitative immunoreactivity (IR) scores and correlated to clinical and pathological data. Different types of galectin expression were compared with respect to disease-free survival (DFS) and overall survival (OS). Gal-8 served as a new positive prognostic factor for the OS and DFS of ovarian cancer patients. Gal-9 expression determined the DFS and OS of ovarian cancer patients in two opposing ways-moderate Gal-9 expression was correlated with a reduced outcome as compared to Gal-9 negative cases, while patients with high Gal-9 expression showed the best outcome.

Glycosylation-dependent binding of galectin-8 to activated leukocyte cell adhesion molecule (ALCAM/CD166) promotes its surface segregation on breast cancer cells.

BACKGROUND: We previously demonstrated that the activated leukocyte cell adhesion molecule (ALCAM/CD166) can interact with galectin-8 (Gal-8) in endothelial cells. ALCAM is a member of the immunoglobulin superfamily that promotes homophilic and heterophilic cell-cell interactions. Gal-8 is a "tandem-repeat"-type galectin, known as a matricellular protein involved in cell adhesion. Here, we analyzed the physical interaction between both molecules in breast cancer cells and the functional relevance of this phenomenon. METHODS: We performed binding assays by surface plasmon resonance to study the interaction between Gal-8 and the recombinant glycosylated ALCAM ectodomain or endogenous ALCAM from MDA-MB-231 breast cancer cells. We also analyzed the binding of ALCAM-silenced or control breast cancer cells to immobilized Gal-8 by SPR. In internalization assays, we evaluated the influence of Gal-8 on ALCAM surface localization. RESULTS: We showed that recombinant glycosylated ALCAM and endogenous ALCAM from breast carcinoma cells physically interacted with Gal-8 in a glycosylation-dependent fashion displaying a differential behavior compared to non-glycosylated ALCAM. Moreover, ALCAM-silenced breast cancer cells exhibited reduced binding to Gal-8 relative to control cells. Importantly, exogenously added Gal-8 provoked ALCAM segregation, probably trapping this adhesion molecule at the surface of breast cancer cells. CONCLUSIONS: Our data indicate that Gal-8 interacts with ALCAM at the surface of breast cancer cells through glycosylation-dependent mechanisms. GENERAL SIGNIFICANCE: A novel heterophilic interaction between ALCAM and Gal-8 is demonstrated here, suggesting its physiologic relevance in the biology of breast cancer cells.