A Newly Identified Horseshoe Crab Lectin with Specificity for Blood Group A Antigen Recognizes Specific O-Antigens of Bacterial Lipopolysaccharides

Abstract

A 14-kDa lectin, named tachylectin-3, was newly identified from hemocytes of the Japanese horseshoe crab, Tachypleus tridentatus. This lectin exhibited hemagglutinating activity against human A-type erythrocytes, but not against the B-and O-types of erythrocytes and animal erythrocytes, including those of sheep, rabbit, horse, and bovine. The hemagglutinating activity of tachylectin-3 was equivalent to that of a previously identified lectin, named tachylectin-2, with affinity for N-acetyl-D-glucosamine or N-acetyl-D-galactosamine. However, the activity of tachylectin-3 was not inhibited by these two N-acetylhexosamines at 100 mM but was inhibited by a blood group A-pentasaccharide at a minimum inhibitory concentration of 0.16 mM. Furthermore, the hemagglutinating activity was strongly inhibited by bacterial S-type lipopolysaccharides (LPSs) from Gramnegative bacteria but not by R-type LPSs lacking Oantigens. One of the most effective S-type LPSs was from Escherichia coli O111:B4, with a minimum inhibitory concentration of 6 ng/ml. These data suggest that tachylectin-3 specifically recognizes Gram-negative bacteria through the unique structural units of O-antigens. Ultracentrifugation analysis revealed that tachylectin-3 is present in dimer in solution. A cDNA coding for tachylectin-3 was isolated from a hemocyte cDNA library. Tachylectin-3 consisted of two repeating sequences, each with a partial sequence similarity to rinderpest virus neuraminidase. Tachylectin-3 and three previously isolated types of tachylectins were all predominantly expressed in hemocytes and released from hemocytes in response to external stimuli. These lectins present at injured sites suggest that they probably serve synergistically to accomplish an effective host defense against invading microbes.Immunity to infectious agents is mediated by two general systems, innate and acquired. Acquired immunity, found only in vertebrates, is one function of B and T lymphocytes, which produce an infinite multitude of specific antigen receptors and antibodies through somatic gene rearrangement. On the other hand, innate immunity is phylogenetically older than acquired immunity, and a certain form of innate immunity is present in all multicellular organisms. Non-self-recognizing proteins involved in innate immunity seem to recognize mainly carbohydrate moieties on pathogens; for instance, a receptor CD14 (1) on macrophages for bacterial lipopolysaccharides (LPSs), 1 Creactive protein (2), and the mannose-binding lectin of the collectin family in mammalian plasma (3). Furthermore, the innate immunity in invertebrates is also triggered by polysaccharides, such as LPSs and ␤-1,3-glucans, as seen in the hemolymph coagulation system in horseshoe crab (4 -9) and the phenoloxidase-mediated melanization system in crustaceans and insects (10).In the Japanese horseshoe crab, Tachypleus tridentatus, one of the major defense systems is carried by hemolymph that contains granular hemocytes comprising 99% of the total hemocytes (11). These granular hemocytes are fil...

Musca domestica Larva Lectin Induces Apoptosis in BEL-7402 Cells through a Mitochondria-Mediated Reactive Oxygen Species Way

“…9) They were ubiquitous in nature, such as in microorganisms, 10) animals 11) and plants. 12) And they played important roles in cellular events like proliferation, 13) antibacterial, 14) signal transduction, 15) and apoptosis. 16) Several lectins have been shown to have antitumor activities.…”

A serine protease zymogen functions as a pattern-recognition receptor for lipopolysaccharides

“…Horseradish peroxidase-conjugated goat anti-mouse IgG (Bio-Rad) was added, and the plates were again incubated at 37°C for 1 h. The enzyme activity of horseradish peroxidase was detected with o-phenylenediamine at 490 nm by using a microplate reader, model 3550 (Bio-Rad). On the other hand, the amount of TL-2 was determined by sandwich ELISA as described (25).…”

A Lipopolysaccharide- and β-1,3-Glucan-binding Protein from Hemocytes of the Freshwater Crayfish Pacifastacus leniusculus

“…Therefore, the proteins involved in the recognition of LPS, peptidoglycans, and ␤-1,3-glucans have been named pattern recognition proteins (20), and they are involved in various ways in the biological defense mechanisms in both invertebrates and vertebrates. Recently, LPS-(21-24) and/or ␤-1,3-glucan-binding proteins (25)(26)(27)(28), peptidoglycan recognition proteins (29 -31), lectins (32)(33)(34), and hemolin (35)(36)(37) have been found in several different species of invertebrates, and their function in the immune response has been studied. For instance, in the horseshoe crab Tachypleus tridentatus, LPS or ␤-1,3-glucans both bind specifically to pattern recognition proteins, and as a result the coagulation cascade is activated (24,28).…”