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M W Johansson

Publications and source records attributed to M W Johansson.

16 recordsLinked to original sources
PubMed2000

A cell adhesion protein from the crayfish Pacifastacus leniusculus, a serine proteinase homologue similar to Drosophila masquerade.

A cDNA encoding a protein resembling masquerade, a serine proteinase homologue expressed during embryogenesis, larval, and pupal development in Drosophila melanogaster, was identified in hemocytes of the adult freshwater crayfish, Pacifastacus leniusculus. The crayfish protein is similar to Drosophila masquerade in the following aspects: (a) overall sequence of the serine proteinase domain, such as the position of three putative disulfide bridges, glycine in the place of the catalytic serine residue, and the presence of a substrate-lining pocket typical for trypsins; (b) the presence of several copies of a disulfide-knotted motif in the putative propeptide. This masquerade-like protein is cleaved into a 27-kDa fragment, which could be detected by immunoblot analysis using an affinity-purified antibody against a synthetic peptide in the C-terminal domain of the protein. The 27-kDa protein could be immunoaffinity-purified from hemocyte lysate supernatant and exhibited cell adhesion activity in vitro, indicating that the C-terminal domain of the crayfish masquerade-like protein mediates cell adhesion.

Amino Acid Sequence↗
PubMed1999

A cell-surface superoxide dismutase is a binding protein for peroxinectin, a cell-adhesive peroxidase in crayfish.

Peroxinectin, a cell-adhesive peroxidase (homologous to human myeloperoxidase), from the crayfish Pacifastacus leniusculus, was shown by immuno-fluorescence to bind to the surface of crayfish blood cells (haemocytes). In order to identify a cell surface receptor for peroxinectin, labelled peroxinectin was incubated with a blot of haemocyte membrane proteins. It was found to specifically bind two bands of 230 and 90 kDa; this binding was decreased in the presence of unlabelled peroxinectin. Purified 230/90 kDa complex also bound peroxinectin in the same assay. In addition, the 230 kDa band binds the crayfish beta-1,3-glucan-binding protein. The 230 kDa band could be reduced to 90 kDa, thus showing that the 230 kDa is a multimer of 90 kDa units. The peroxinectin-binding protein was cloned from a haemocyte cDNA library, using immuno-screening or polymerase chain reaction based on partial amino acid sequence of the purified protein. It has a signal sequence, a domain homologous to CuZn-containing superoxide dismutases, and a basic, proline-rich, C-terminal tail, but no membrane-spanning segment. In accordance, the 90 and 230 kDa bands had superoxide dismutase activity. Immuno-fluorescence of non-permeabilized haemocytes with affinity-purified antibodies confirmed that the crayfish CuZn-superoxide dismutase is localized at the cell surface; it could be released from the membrane with high salt. It was thus concluded that the peroxinectin-binding protein is an extracellular SOD (EC-SOD) and a peripheral membrane protein, presumably kept at the cell surface via ionic interaction with its C-terminal region. This interaction with a peroxidase seems to be a novel function for an SOD. The binding of the cell surface SOD to the cell-adhesive/opsonic peroxinectin may mediate, or regulate, cell adhesion and phagocytosis; it may also be important for efficient localized production of microbicidal substances.

Amino Acid Sequence↗
PubMed1997

Identification and cloning of an integrin beta subunit from hemocytes of the freshwater crayfish Pacifastacus leniusculus.

We have cloned and sequenced a beta subunit of integrin from a cDNA library of crayfish hemocytes. This beta integrin shows great similarity to beta integrin subunits from other animals; the highest is towards beta pat-3 from Caenorhabditis elegans followed by beta PS from Drosophila melanogaster. By immunoblotting with antibodies raised towards a synthetic peptide corresponding to a part of the cytoplasmic region of the deduced protein sequence, it was shown that the integrin is present in the membrane of the hemocytes. This is the first integrin found in hemocytes of an invertebrate animal and this finding opens the door for further investigations on integrins and their role in the invertebrate immune system.

Amino Acid Sequence↗
PubMed1997

Myeloperoxidase mediates cell adhesion via the alpha M beta 2 integrin (Mac-1, CD11b/CD18).

Myeloperoxidase is a leukocyte component able to generate potent microbicidal substances. A homologous invertebrate blood cell protein, peroxinectin, is not only a peroxidase but also a cell adhesion ligand. We demonstrate in this study that human myeloperoxidase also mediates cell adhesion. Both the human myeloid cell line HL-60, when differentiated by treatment with 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or retinoic acid, and human blood leukocytes, adhered to myeloperoxidase; however, undifferentiated HL-60 cells showed only minimal adhesion. No cells adhered to horseradish peroxidase, and cell adhesion to myeloperoxidase was not decreased by catalase, thus showing that peroxidase activity, per se, was neither sufficient nor necessary for the adhesion activity. Mannan, which has been reported to inhibit the binding of peroxidases to cells, did not affect adhesion to myeloperoxidase. However, adhesion to myeloperoxidase was inhibited by monoclonal antibodies to alpha M (CD11b) or to beta2 (CD18) integrin subunits, but not by antibodies to alpha L (CD11a), alpha M (CD11c), or to other integrins. Native myeloperoxidase mediated dose-dependent cell adhesion down to relatively low concentrations, and denaturation abolished the adhesion activity. It is evident that myeloperoxidase supports cell adhesion, a function which may be of considerable importance for leukocyte migration and infiltration in inflammatory reactions, that alpha M beta2 integrin (Mac-1 or CD11b/CD18) mediates this adhesion, and that the alphaM beta2 integrin-mediated adhesion to myeloperoxidase is distinct from the previously reported ability of this integrin to bind to certain denatured proteins at high concentrations.

Antibodies, Monoclonal↗
PubMed1996

The prophenoloxidase activating system and associated proteins in invertebrates.

In this review, we present arguments indicating that prophenoloxidase (proPO) activating system acts as a pattern recognition and defence system in invertebrate blood. Phenoloxidase (PO) activity has been found in the blood of many invertebrates. At least in arthropods, echinoderms and urochordates, the inactive pro-form, proPO has been found to be elicited by the microbial cell-wall components beta-1, 3-glucans, lipopolysaccharide and/or peptidoglycan. This activation seems to involve elicitor-binding proteins and serine protease(s). ProPO, the proPO-activating enzyme (ppA) and plasma elicitor-binding proteins, have been purified from some arthropods, and proPO and the beta-1, 3-glucan binding protein (beta GBP) have been cloned and sequenced from crayfish. Arthropod proPO has a molecular mass of 70-90 kDa and PO has a molecular mass of 60-70 kDa. The beta GBP also stimulates phagocytosis of fungal cells and, after reacting with beta-1, 3-glucan, blood-cell degranulation (and release of the proPO system). In addition, a cell-adhesion protein (of 70-100 kDa), apparently associated with the proPO system, has been purified from arthropods. This mediates blood-cell adhesion, degranulation, phagocytosis and encapsulation. The cell-adhesion protein and beta GBP bind to a common blood-cell membrane receptor. It would be interesting to see the sequences of more proPO system components and investigate whether the scheme for cellular communication and defence, involving the cell-adhesion protein, elicitor-binding proteins and the membrane receptor described in arthropods, applies to invertebrates in general.

Animals↗
PubMed1995

Peroxinectin, a novel cell adhesion protein from crayfish blood.

From blood cells of the crayfish Pacifastacus leniusculus a 76-kDa protein that mediated attachment and spreading of the crayfish blood cells was purified. The cDNA for this cell adhesion protein was isolated, cloned, and sequenced. The deduced protein sequence was significantly similar to one family of peroxidases, e.g., myeloperoxidase. Consistently, the 76-kDa protein, for which we propose the name peroxinectin, had peroxidase activity. A synthetic peptide derived from the peroxinectin sequence containing Lys-Gly-Asp mimicked the cellular activity of the intact protein, implicating this sequence as the cell-binding site. Peroxinectin is the first cell adhesion molecule cloned from invertebrate blood and, to our knowledge, the first protein from any organism that combines being a cell adhesion ligand and a peroxidase.

Amino Acid Sequence↗
PubMed1994

Purification and cDNA cloning of a four-domain Kazal proteinase inhibitor from crayfish blood cells.

A cDNA with an open reading frame of 684 base pairs was isolated from a library from blood cells of the crayfish Pacifastacus leniusculus. It codes for a signal sequence and a mature protein of 209 amino acids with a predicted molecular mass of 22.7 kDa. The amino acid sequence consists of four repeated stretches (45-73% identical to each other), indicating that the protein has four domains. The domains have significant sequence similarity to serine proteinase inhibitors of the Kazal family. The three first domains have a leucine residue in the putative reactive site, suggesting that the protein is a chymotrypsin inhibitor. A monomeric 23-kDa proteinase inhibitor, which by amino terminal sequencing of the mature protein was confirmed to be the cloned Kazal inhibitor, was purified from crayfish blood cells. It inhibited chymotrypsin or subtilisin, but not trypsin, elastase or thrombin. The inhibitor seemed to form a 1:1 complex with chymotrypsin or subtilisin. This protein seems to be the first described Kazal inhibitor from blood cells of any animal and the first one with four domains.

Amino Acid Sequence↗
PubMed1994

Altered localization and cytoplasmic domain-binding properties of tyrosine-phosphorylated beta 1 integrin.

We describe a novel approach to study tyrosine-phosphorylated (PY) integrins in cells transformed by virally encoded tyrosine kinases. We have synthesized a peptide (PY beta 1 peptide) that represents a portion of the cytoplasmic domain of the beta 1 integrin subunit and is phosphorylated on the tyrosine residue known to be the target of oncogenic tyrosine kinases. Antibodies prepared against the PY beta 1 peptide, after removal of cross-reacting antibodies by absorption and affinity purification, recognized the PY beta 1 peptide and the tyrosine-phosphorylated form of the intact beta 1 subunit, but did not bind the nonphosphorylated beta 1 peptide, the nonphosphorylated beta 1 subunit or other unrelated tyrosine-phosphorylated proteins. The anti-PY beta 1 antibodies labeled the podosomes of Rous sarcoma virus-transformed fibroblasts, but did not detectably stain nontransformed fibroblasts. The localization of the tyrosine phosphorylated beta 1 subunits appeared distinct from that of the beta 1 subunit. Adhesion plaques were stained by the anti-beta 1 subunit antibodies in Rous sarcoma virus-transformed fibroblasts plated on fibronectin, whereas neither podosomes nor adhesion plaques were labeled on vitronectin or on uncoated plates. Anti-phosphotyrosine antibodies labeled podosomes, adhesion plaques and cell-cell boundaries regardless of the substratum. One of the SH2 domains of the p85 subunit of phosphatidylinositol-3-kinase bound to the PY beta 1 peptide, but not to the non-phosphorylated beta 1 cytoplasmic peptide. Other SH2 domains did not bind to the PY beta 1 peptide. These results show that the phosphorylated form of the beta 1 integrin subunit is detected in a different subcellular localization than the nonphosphorylated form and suggest that the phosphorylation on tyrosine of the beta 1 subunit cytoplasmic domain may affect cellular signaling pathways.

3T3 Cells↗
PubMed1989

Cellular immunity in crustaceans and the proPO system.

The molecular mechanism of cellular immunity in arthropods has until recently been largely unknown, but with the development of a technique to isolate and handle the different blood cell types of crustaceans and with the purification of several proteins associated with the so-called proPO system of freshwater crayfish the processes have now begun to be better understood. In this article Mats Johansson and Kenneth Söderhäll discuss the function of the proPO system in cellular immune reactions in crustaceans and in particular the role of a protein with a molecular mass of 76 kDa, which has been shown to be involved in the communication between the different blood cell types of crayfish.

Journal Article↗
PubMed1988

Isolation and purification of a cell adhesion factor from crayfish blood cells.

Isolated granular haemocytes (blood cells) from the crayfish Pacifastacus leniusculus attached and spread in vitro on coverslips coated with a lysate of crayfish haemocytes. No cell adhesion activity was detected in crayfish plasma. The cell adhesion activity was only present in haemocyte lysates in which the prophenoloxidase (proPO) activating system (Söderhäll and Smith, 1986a, b) had been activated; either by lipopolysaccharide (LPS), the beta-1,3-glucan laminarin, or by preparing the lysate in 5 mM Ca2+. Both lysates of granular or of semigranular haemocytes could mediate adhesion. After A23187-induced exocytosis of the granular cells, cell adhesion activity could be generated in the secreted material if it was incubated with laminarin. The factor responsible for cell adhesion was isolated from an active haemocyte lysate and purified by ammonium sulfate precipitation, cation exchange chromatography and Con A-Sepharose; it had a molecular mass of approximately 76 kD on an SDS-polyacrylamide gel. An antibody to this 76-kD band inhibited cell adhesion. Ca2+ was necessary in the medium for the cells to adhere to the adhesion factor. With cyanide or azide, the cells attached but failed to spread. It is suggested that in vivo the cell adhesion factor is stored in the secretory granules of the semigranular and the granular cells in a putative inactive pro-form, which can be released during exocytosis and, in the presence of beta-1,3-glucans or LPS, be activated outside the cells to mediate cell attachment and spreading, processes of essential importance in arthropod host defense.

Animals↗
PubMed1985

The cytotoxic reaction of hemocytes from the freshwater crayfish, Astacus astacus.

Crayfish hemocytes displayed cytotoxic capacity towards all tested mammalian tumor and nontumor cell lines. The ratio required for the cytotoxic action of effector cells to target cells was at least 1:1. The lysis of the target cells required a minimum of 1 hr to become detected. After separation and isolation of the hemocyte populations of crayfish, the semigranular and granular cells retained their cytotoxic capacity. These cells contain the prophenoloxidase activating (proPO) system, a complement-like pathway, which in an activated form lyses semigranular cells in vitro, but failed to kill the tested target cells.

Animals↗
PubMedDate not supplied

Opsonic activity of cell adhesion proteins and beta-1,3-glucan binding proteins from two crustaceans.

A beta-1,3-glucan binding protein (beta GBP) from the shore crab Carcinus maenas was purified from plasma by precipitation of the protein at low ionic strength. The protein had a molecular mass of 110 kDa, and was shown to affinity precipitate with laminarin, a soluble beta-1,3-glucan, and to cross-react with an antiserum directed toward beta GBP from the crayfish Pacifastacus leniusculus. Also, a protein from the haemocytes of C. maenas with a molecular mass of 80 kDa was found to mediate cell attachment and cause degranulation of crab cells, similar to the 76 kDa protein present in the haemocytes of P. leniusculus. Antibodies against the crayfish 76-kDa protein reacted with the crab 80-kDa protein present in the granular cells. No 80-kDa protein could be found in the hyaline cells. Using a method with FITC-conjugated yeast particles in a phagocytosis assay, both the beta GBP and the 80-kDa protein from C. maenas were shown to have opsonic activity as had beta GBP and 76-kDa protein from P. leniusculus, resulting in higher levels of phagocytosis by the crab hyaline cells. Treatment of the yeast particles with beta GBP previously reacted with laminarin (beta GBP-L) only resulted in a minor increase of phagocytosis. Moreover, if the phagocytic cells were preincubated with beta GBP-L or with the 80-kDa protein, the enhancement of the phagocytic activity by beta GBP or the 80-kDa protein were abolished, indicating that a saturable number of one kind of cell surface receptor seem to be involved in phagocytosis.

Animals↗
PubMedDate not supplied

Peroxinectin, a cell adhesive protein associated with the proPO system from the black tiger shrimp, Penaeus monodon.

Upon activation of the prophenoloxidase activating system in the shrimp, Penaeus monodon, a cell adhesion activity in the haemolymph is generated. A cell adhesion assay showed that a high number of granular cells (60%) adhered to coverslips coated with a shrimp haemocyte lysate supernatant, whereas a very low number of cells adhered to coverslips coated with bovine serum albumin. Inhibition of adhesion by an antiserum against crayfish peroxinectin, a cell adhesion protein, revealed that the cell adhesion activity detected in shrimp haemocyte lysate supernatant might result from a peroxinectin-like molecule in shrimp. A cDNA clone encoding shrimp peroxinectin was isolated, which had an open reading frame of 2337 nucleotides, with a polyadenylation sequence and a poly A tail. It encodes a protein of 778 amino acids including a 20 amino acid signal peptide. The mature protein (758 amino acids) has a predicted molecular mass of 84.8kDa and an estimated pI of 9.0. Two putative integrin binding motifs, RGD (Arg-Gly-Asp) and KGD (Lys-Gly-Asp), were found in shrimp peroxinectin. Sequence comparison shows that the shrimp protein is similar to crayfish peroxinectin (69%) and to various peroxidases and putative peroxidases from invertebrates and vertebrates. The shrimp peroxinectin cDNA also shows similarity (51%) to both Drosophila peroxinectin-related protein (AAF78217) and peroxidasin (S46224), an extracellular matrix protein combining an active peroxidase domain as well as immunoglobulin domains, leucine rich repeats and procollagen-like motif. However, the sequence similarity to both Drosophila molecules are mostly within the peroxidase domain. Northern blot analysis, using a non-peroxidase region in peroxinectin as a probe, revealed that peroxinectin is constitutively expressed in shrimp haemocyte and was reduced significantly in shrimp injected with a beta-1,3-glucan, laminarin, to mimic an infection with a fungus.

Amino Acid Sequence↗
PubMedDate not supplied

Intracellular signaling in arthropod blood cells: involvement of protein kinase C and protein tyrosine phosphorylation in the response to the 76-kDa protein or the beta-1,3-glucan-binding protein in crayfish.

Degranulation (regulated exocytosis) of crayfish granular blood cells and release of the prophenoloxidase activating system can be triggered by two endogenous ligands, a 76-kDa cell adhesion protein or a beta-1,3-glucan-binding protein when reacted with beta-1,3-glucan (beta GBP-L). These ligands bind a recently described membrane receptor. Degranulation triggered by these ligands was inhibited by the protein kinase C (pkC) inhibitor staurosporine or by the tyrosine kinase inhibitor herbimycin A. Incubation with the 76-kDa protein increased tyrosine phosphorylation of a granular cell protein of approximately 80 kDa. In addition, the pkC activating phorbol esters PMA or beta PDD degranulated the cells in a dose-dependent manner, whereas the control isomeric phorbol ester alpha PDD that does not activate pkC did not have any effect on the cells. Thus, we propose that binding of the 76-kDa protein or beta GBP-L to the receptor triggers cellular responses via a pathway that includes pkC activation and protein tyrosine phosphorylation.

Alkaloids↗
PubMedDate not supplied

Cell adhesion molecules in invertebrate immunity.

Cell adhesion is essential in immunity in invertebrates, e.g., in the cellular immune responses of encapsulation and nodule formation. Here cell adhesion molecules shown or suggested to be involved in invertebrate immunity are reviewed. Blood cells of the crayfish, Pacifastacus leniusculus, can release a cell-adhesive and opsonic peroxidase, peroxinectin. A site containing the motif, KGD, appears to be adhesive by binding to a transmembrane receptor of the integrin family on the blood cells. Peroxinectin also binds a peripheral blood cell surface CuZn-superoxide dismutase. The peroxidase-integrin interaction appears to have evolved early and seems conserved; human myeloperoxidase supports cell adhesion via the alphaMbeta2 integrin. There is evidence for peroxinectin-like proteins in other arthropods. Effects by RGD peptides indicate that integrins mediate blood cell adhesion and cellular immunity in diverse invertebrate species. Other invertebrate blood cell molecules proposed to be involved in adhesion include the insect plasmatocyte-spreading peptide, as well as soluble and transmembrane proteins which show some similarity to vertebrate adhesive or extracellular matrix molecules. Proteins such as the Ig family member hemolin, or proteins found in insects that are hosts for parasitic wasps, inhibit cell adhesion and may regulate or block cellular immunity.

Animals↗