PubMed Health⌕ Search

Biomedical subjects

M E Perkins

Publications and source records attributed to M E Perkins.

At least 37 records · Page 2Linked to original sources
PubMed1989

Localization of a parasite encoded protein to erythrocyte cytoplasmic vesicles of Plasmodium falciparum-infected cells.

Intracellular development of the malarial parasite results in substantial modifications of the membrane and cytoskeleton of the erythrocyte host cell. Two related Plasmodium falciparum-encoded proteins of 50 kDa and 43 kDa (Pf 50/43), identified by reactivity with a single mAb, were demonstrated to be localized to the erythrocyte cytoplasm of parasite-infected cells. Immunofluorescence and immunoelectron microscopy using mAb.7E11 demonstrated the Pf 50/43 is localized in the membrane of the vesicles in the erythrocyte cytoplasm, vesicles which correspond to Maurer's clefts. Solubility properties of the proteins suggest they are integral membrane proteins. By immunofluorescence, Pf 50/43 is shown to colocalize with actin which has a highly modified organization in the infected erythrocyte. Pf 50/43 is located exclusively in the vesicles, is not transported to the erythrocyte membrane or secreted. It is proposed the vesicles may play a role in transport of molecules across the erythrocyte cytoplasm, between the parasite and the external erythrocyte plasma membrane.

Animals↗
PubMed1988

Sialic acid-dependent binding of Plasmodium falciparum merozoite surface antigen, Pf200, to human erythrocytes.

Plasmodium falciparum merozoites, the extracellular stage of the erythrocytic cycle of the human malarial parasite, specifically invade human E. The major determinant of that specificity is the sialic acid residues of E glycophorin. In the present study we show that the merozoite surface Ag, Pf200 (m.w. 195,000 to 205,000), of two different isolates of P. falciparum, binds to the surface of human E but not E from other species not invaded by P. falciparum. Pf200 does not bind to neuraminidase-treated E, indicating the interaction is dependent on sialic acid residues. Binding is inhibited by soluble glycophorin and selective mAb against the glycosylated domain of glycophorin, but not by a mAb against the peptide domain of glycophorin. mAb.5B1 previously identified as reacting with Pf 200, blocks binding of the protein to the E. Binding between Pf200 and the E is not high affinity, as Pf200 can be released from the surface by 0.25 M NaCl.

Animals↗
PubMed1988

Erythrocyte receptor recognition varies in Plasmodium falciparum isolates.

N-Acetylneuraminic acid (NeuNAc) is the terminal sugar residue of the O-linked tetrasaccharide linked to erythrocyte sialoglycoproteins, glycophorins. Erythrocytes lacking NeuNAc have been shown previously to be resistant to invasion by certain isolates of Plasmodium falciparum merozoites. We report here variation between different geographic isolates of P. falciparum in their dependency on NeuNAc for invasion of host erythrocytes. Seven different geographic isolates of P. falciparum were examined for their ability to invade neuraminidase treated erythrocytes. For all isolates invasion was reduced significantly, although considerable variation in NeuNAc dependency was apparent. Three isolates, FCR-3, FVO and It2, exhibited a very high dependence on NeuNAc residues for invasion (invasion reduced greater than 90%), whereas two isolates (Thai-Tn and FC-27) exhibited a moderately high dependence (invasion reduced 75%). Two other isolates (CDC-1 and 7G8) exhibited moderate dependence on NeuNAc (invasion reduced 50%). Cleavage of the complete O-linked tetrasaccharide by O-glycanase removes all carbohydrate from glycophorin A, B and C except the single N-linked oligosaccharide on glycophorin A and C. Invasion of FCR-3 and CDC-1 isolates into O-glycanase treated erythrocytes was not markedly different from that into neuraminidase treated cells indicating that NeuNAc is the important residue of the tetrasaccharide for both isolates. Invasion into endo-beta-galactosidase treated erythrocytes, in which the lactosaminoglycan side chain of band 3 and band 4.5 is cleaved, was not significantly reduced for either the CDC-1 or FCR-3 isolates. Additional results on the trypsin insensitivity of band 3 also suggest that this erythrocyte protein is not important in P. falciparum recognition. The greatest divergence in receptor specificity between FCR-3 and CDC-1 isolates was apparent in invasion into periodate-treated erythrocytes. Periodate oxidation results in cleavage of the exocyclic hydroxyl groups of the terminal NeuNAc but leaves its COOH group unaltered. These experiments also illustrated that the negatively charged COOH group of NeuNAc is not the important group in the interaction of the merozoite with the NeuNAc. Trypsin-treated erythrocytes were almost fully resistant to invasion by CDC-1 as well as the FCR-3 isolates suggesting that the CDC-1 isolate, in addition to interacting with NeuNAc, depends on a trypsin sensitive site for invasion. This site could involve the N-linked saccharide on glycophorin A and C or a protein on the erythrocyte surface unrelated to the glycophorins.

Animals↗
PubMed1988

Secretion of Plasmodium falciparum rhoptry protein into the plasma membrane of host erythrocytes.

The rhoptry is an organelle of the malarial merozoite which has been suggested to play a role in parasite invasion of its host cell, the erythrocyte. A monoclonal antibody selected for reactivity with this organelle identifies a parasite synthesized protein of 110 kD. From biosynthetic labeling experiments it was demonstrated that the protein is synthesized midway through the erythrocytic cycle (the trophozoite stage) but immunofluorescence indicates the protein is not localized in the organelle until the final stage (segmenter stage) of intraerythrocytic development. Immunoelectron microscopy shows that the protein is localized in the matrix of the rhoptry organelle and on membranous whorls secreted from the merozoite. mAb recognition of the protein is dithiothreitol (DTT) labile, indicating that the conformation of the epitope is dependent on a disulfide linkage. During erythrocyte reinvasion by the extracellular merozoite, immunofluorescence shows the rhoptry protein discharging from the merozoite and spreading around the surface of the erythrocyte. The protein is located in the plasma membrane of the newly invaded erythrocyte. These studies suggest that the 110-kD rhoptry protein is inserted into the membrane of the host erythrocyte during merozoite invasion.

Animals↗
PubMed1985

Wright (a + b-) human erythrocytes and Plasmodium falciparum malaria.

We find Wr(a + b-) erythrocytes of donor M. Fr., which appear to carry a rare glycophorin A variant, to be fully susceptible to invasion by nine isolates of Plasmodium falciparum. Thus we fail to confirm the previous publication on the refractoriness of these erythrocytes. In addition the serum of donor M. Fr., which is known to contain anti-Wrb directed against an epitope located on glycophorin A in close proximity to the erythrocyte membrane, was not found to inhibit P. falciparum invasion of blood group O Rh- red blood cells. Despite this, different lines of evidence still indicate that glycophorin A is one of the receptors for erythrocyte invasion by P. falciparum. The Wrb epitope, however, does not appear to represent a distinct receptor site, which is in contrast to previous suggestions.

ABO Blood-Group System↗
PubMed1985

Characterization with monoclonal antibodies of a surface antigen of Plasmodium falciparum merozoites.

The merozoite, the extracellular form of the erythrocyte stage of the malarial parasite, invades the erythrocyte and develops intracellularly. Cloned hybridoma cell lines secreting monoclonal antibodies directed against the merozoite surface were selected by indirect immunofluorescent assay by using intact isolated merozoites. Monoclonal antibodies to a 200,000 m.w. merozoite surface antigen were selected and were used to characterize this protein and its role in erythrocyte invasion. Immunoelectron microscopy demonstrated that the antigen was located exclusively on the merozoite surface coat, distributed evenly over the entire surface. The 200,000 m.w. protein incorporated [3H]glucosamine, suggesting that it is a glycoprotein and could be purified to homogeneity by using immuno-affinity chromatography. Freshly isolated, invasive merozoites retained the 200,000 m.w. antigen, but the protein was rapidly cleaved to proteins of 90,000 and 50,000 m.w. when the merozoite was extracellular. The 50,000 m.w. fragment was retained the epitope binding to monoclonal antibody 5B1 and were labeled with [3H]glucosamine. Monoclonal antibodies against the 200,000 m.w. antigen partially inhibited merozoite invasion into erythrocytes.

Animals↗
PubMed1984

Surface proteins of Plasmodium falciparum merozoites binding to the erythrocyte receptor, glycophorin.

Invasion of erythrocytes by the malarial parasite is a receptor-mediated process. P. falciparum merozoites recognize and bind to erythrocyte surface sialoglycoproteins, glycophorins A and B, and the glycophorins bind to saturable sites on the merozoite surface. The purpose of the present work was to identify a receptor or ligand molecule on the merozoite surface that mediates binding to the erythrocyte. A fraction containing the sialoglycoproteins was coupled to an acrylamide matrix and incubated with metabolically labeled merozoites. A merozoite protein of 155 kD that labeled prominently with [3H]glycine bound to glycophorin. A minor protein of 130 kD also bound. Both proteins are rich in proline and glycine, poor in methionine, and may be related. The proteins are also stable to heating to 100 degrees C for 10 min. Immunoelectron microscopy demonstrated that the 155 kD and 130 kD proteins are located on the merozoite surface coat. The antibodies significantly inhibited merozoite invasion into erythrocytes and also binding of the proteins to the glycophorin-matrix. The specific binding of the 155-kD and 130-kD proteins to the erythrocyte receptor and the demonstration that they are located on the merozoite surface suggest they could be receptor proteins that mediate binding of the merozoite to the erythrocyte surface.

Animals↗
PubMed1984

Binding of glycophorins to Plasmodium falciparum merozoites.

Plasmodium falciparum merozoites recognize and attach to glycophorins, the surface sialoglycoproteins of human erythrocytes. The structural requirements for a merozoite binding site were studied with the use of two methods. In the first, certain glycophorins and their tryptic fragments were added directly to isolated merozoites prior to their addition to erythrocytes. Low concentrations (50 micrograms ml-1) of glycophorin A inhibited merozoite invasion. At higher concentrations a mixture of glycophorins A, B and C (GPS) (100 micrograms ml-1) and glycophorin B (200 micrograms ml-1) also inhibited invasion. GPS from Tn erythrocytes which lack both sialic acid and galactose residues was almost as effective as normal GPS in blocking invasion. None of the monosaccharides present on glycophorin, including N-acetylneuraminic acid, inhibited merozoite invasion. Erythrocytes treated with lectins were only partially resistant to invasion. These results indicated that the oligosaccharide side chains are not the major structural determinant of the merozoite binding site. Glycophorin A was cleaved by trypsin and the separated fragments added to merozoites. Only the external N-terminal tryptic fragment T1 and the trypsin resistant hydrophobic core, T6, showed some, but considerably less, inhibitory activity than the intact molecule. In the second approach, the binding of 125I-labeled GPS to isolated merozoites was determined. 125I-GPS binding was saturated at 0.23 micrograms for 10(9) merozoites and was competitively inhibited by unlabeled GPS but not by free sugars. Desialylated GPS bound almost to the same extent as the intact molecule.

Animals↗
PubMed1984

Receptor mediated endocytosis of the malarial parasite by erythrocytes.

Malarial merozoites appear to interact with the major surface sialoglycoproteins, glycophorin A and glycophorin B, during entry into the human erythrocyte. Intact merozoites bind 125I-glycophorin demonstrating directly that the parasites have glycophorin binding sites. A glycophorin containing fraction crosslinked to acrylamide beads provided a probe to identify merozoite molecules with an affinity for the erythrocyte surface. A fraction of merozoites, labeled with 3H-proline, and containing the membrane proteins and a fraction containing soluble proteins released by the merozoite were added to the glycophorin beads. Two proteins, M.W. of 155,000 and 130,000, specifically bound to glycophorin. These proteins may represent the merozoite recognition site for the erythrocyte.

Animals↗
PubMed1979

Cell surface fibronectin and oncogenic transformation.

Fibronectin is a large glycoprotein at the cell surface of many different cell types; a related protein is present in plasma. Fibronectin is a dimer of 230,000-dalton subunits and also occurs in larger aggregates; it forms fibrillar networks at the cell surface, between cells and substrata and between adjacent cells, and it is not a typical membrane protein. Cell surface fibronectin is reduced in amount or absent on transformed cells and in many cases its loss correlates with acquisition of tumorigenicity and, in particular, metastatic ability. Exceptions to the correlations with transformation and tumorigenicity exist. Loss of fibronectin and the resulting reduced adhesion appear to be involved in pleiotropic alterations in cell behavior and may be responsible for several aspects of the transformed phenotype in vitro. Fibronectin interacts with other macromolecules (collagen/gelatin, fibrin/fibrinogen, proteoglycans) and is apparently connected to microfilaments inside the cell.

Animals↗
PubMed1979

Cross-linking of fibronectin to sulfated proteoglycans at the cell surface.

Fibronectin is a major surface protein of normal animal cells but is absent from many transformed cells. Addition of fibronectin to transformed cells causes increased cell substrate adhesion and changes in the morphology and cytoskeleton of the cells. We have coupled fibronectin to photoactivable chemical cross-linkers and have added it to cells to identify those molecules to which it binds. In this way, fibronectin can be cross-linked to sulfated proteoglycans at the cell surface. The cross-linking is specific for fibronectin. The fibronectin-proteoglycan complex is sensitive to chondroitinase ABC and AC and to trypsin. Addition of fibronectin also affects binding of hyaluronic acid to the cells. These results suggest that fibronectin interacts with proteoglycans at the cell surface. The existence of such interactions may have implications for the role of fibronectin and proteoglycans in cell adhesion.

Azides↗
PubMed1979

Autoshaping with common and distinctive stimulus elements, compact and dispersed arrays.

Four groups of pigeons were trained with a standard autoshaping procedure in which a brief fixed-duration interval always followed by a grain delivery alternated with a longer variable-duration interval never associated with grain delivery. One of two stimuli was always presented during each interval. One of them contained three black dots and a black star on a green background; the other contained four black dots on a green background. The four elements of each stimulus were arranged in a more compact array for two groups and in a more dispersed array for the other two groups. Which of the two stimuli preceded grain delivery was counterbalanced within each pair of groups. The speed of occurrence of the first autoshaped peck was not affected by whether the stimulus containing the distinctive star element preceded grain delivery, but autoshaping was faster when the stimulus arrays were compact than when they were dispersed. During 560 response-independent training trials that followed the first autoshaped peck, this pattern reversed; both discriminative control over responding and the relative frequency of pecking the stimulus that preceded grain delivery were greater for the two groups where this stimulus contained the discriminative element than for the two groups where it contained only common elements. During subsequent testing with stimuli containing only a single element each, the distinctive feature was responded to proportionately more often by the two groups for which it had been an element of the stimulus preceding grain delivery than by the two groups for which it had been an element of the stimulus complex that never was associated with grain delivery. These data add further support to the hypothesis that the initial occurrence of autoshaped responding and its subsequent maintenance are not affected by the same variables. They also suggest that automaintenance is as sensitive as response-dependent training to the presence or absence of a distinctive stimulus element among several common elements and that this sensitivity appears to be independent of the specific method used for presenting the stimuli during automaintenance.

Journal Article↗
PubMed1977

Stimulus generalization from feeder to response key in the acquisition of autoshaped pecking.

During autoshaping, a 6-second presentation of one stimulus and a variable time 30-second presentation of a second stimulus alternated in appearance on a pigeon key. Grain always was delivered for 3 seconds at the end of the first stimulus interval. In the first experiment, autoshaped pecking of the stimulus preceding grain delivery began much sooner when that stimulus was a black vertical line on a white background and the other stimulus was green than when the opposite stimulus arrangement was used. Because these two stimuli differed in form, hue, brightness, and similarity in hue and brightness to the illumination of the raised feeder, three subsequent experiments examined whether the differential speed of autoshaping in the two groups was due to a feature-positive, feature-negative effect, a preference for brighter over darker stimuli, a simple preference for white over green, or stimulus generalization from the brightness or hue of the illuminated, raised feeder to the stimulus on the key preceding grain delivery. The data from these experiments showed that the first autoshaped key peck was most likely to be made to the stimulus of the same hue as that illuminating the feeder, regardless of whether that stimulus was positively or negatively associated with grain delivery. At least under some conditions, therefore, stimulus-generalization mediated response transfer of pecking grain in the presence of the hue illuminating the feeder to pecking the key illuminated by a similar hue appears to account for the occurrence of autoshaped key pecking.

Journal Article↗