Physical mapping and expression of gene families encoding the <i>N</i>‐acetyl <scp>d</scp>‐galactosamine adherence lectin of <i>Entamoeba histolytica</i>

Ramakrishnan, Girija; Ragland, Brian D.; Purdy, Jay E.; Mann, Barbara J.

doi:10.1046/j.1365-2958.1996.356885.x

Cited by 47 publications

(27 citation statements)

References 28 publications

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“…3). The hgl probe hybridized with five to seven bands, depending on the strain, as has been previously reported (35).…”

Section: Vol 69 2001supporting

confidence: 78%

“…Clamped homogenous electric field (CHEF) gels of genomic DNA digested with HindIII were electrophoresed in a Bio-Rad CHEF DRIII apparatus as described previously (35). CHEF gels were dried down and used directly in hybridization.…”

Section: Vol 69 2001mentioning

confidence: 99%

“…It was originally identified by galactose affinity chromatography and with adherence-inhibitory monoclonal antibodies (MAbs) (30,43). Both Hgl and Lgl are encoded by gene families (28,35). Antibodies that block or augment parasite Gal/GalNAc binding activity map to the cysteine-rich region (amino acids 356 to 1143) of Hgl (25), and this region (when expressed in Escherichia coli) contains a functional carbohydrate recognition domain (14, 33).…”

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confidence: 99%

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Intermediate Subunit of the Gal/GalNAc Lectin of Entamoeba histolytica Is a Member of a Gene Family Containing Multiple CXXC Sequence Motifs

et al. 2001

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Killing by Entamoeba histolytica requires parasite adherence to host galactose-and N-acetyl-D-galactosamine (Gal/GalNAc)-containing cell surface receptors. A 260-kDa heterodimeric E. histolytica Gal/GalNAc lectin composed of heavy (Hgl) and light (Lgl) subunits has been previously described. Here we present the cloning and characterization of Igl, a 150-kDa intermediate subunit of the Gal/GalNAc lectin. Igl, Hgl, and Lgl colocalized on the surface membrane of trophozoites. Two unlinked copies of genes encoding Igl shared 81% amino acid sequence identity (GenBank accession no. AF337950 and AF337951). They encoded cysteine-rich proteins with amino-and carboxy-terminal hydrophobic signal sequences characteristic of glycosylphosphatidylinositol (GPI)-anchored membrane proteins. The igl genes lacked carbohydrate recognition domains but were members of a large family of amebic genes containing CXXC and CXC motifs. These data indicate that Igl is part of the parasite's multimolecular Gal/GalNAc adhesin required for host interaction.Carbohydrate-protein interactions initiate the contact-dependent cytotoxicity for which Entamoeba histolytica was named. Parasite recognition of host galactose (Gal) and Nacetyl-D-galactosamine (GalNAc) residues initiates trophozoite adherence to human colonic mucin, colonic epithelium, neutrophils and erythrocytes, certain bacteria, and a variety of cultured cell lines (3-7, 16, 19-22, 27, 36-38). Contact-dependent killing of target cells is Ͼ90% inhibited by Gal and GalNAc (34,37,41). Additionally, Chinese hamster ovary (CHO) cell glycosylation-deficient mutants lacking terminal Gal/GalNAc residues on N-and O-linked sugars are nearly totally resistant to amebic adherence and cytolytic activity (23,24,39).The E. histolytica 260-kDa Gal/GalNAc lectin is a heterodimer of transmembrane heavy (170 kDa) (Hgl) and GPIanchored light (35 or 31 kDa) (Lgl) glycoproteins linked by disulfide bonds. It was originally identified by galactose affinity chromatography and with adherence-inhibitory monoclonal antibodies (MAbs) (30,43). Both Hgl and Lgl are encoded by gene families (28,35). Antibodies that block or augment parasite Gal/GalNAc binding activity map to the cysteine-rich region (amino acids 356 to 1143) of Hgl (25), and this region (when expressed in Escherichia coli) contains a functional carbohydrate recognition domain (14, 33). The cytoplasmic tail of Hgl has homology to the cytoplasmic domain of ␤2 and ␤7 integrins, including regions implicated in binding of the intracellular signaling molecules Shc and Grb2. Overexpression of the cytoplasmic tail results in a dominant negative effect on endogenous lectin activity, with decreased adherence, cytotoxicity, and in vivo virulence (44).The 150-kDa lectin intermediate subunit (Igl) was originally identified as a trophozoite surface antigen recognized by MAbs which block trophozoite adherence to mammalian cells in vitro (9)(10)(11)42). The EH3015 MAb specific for Igl significantly inhibits adherence of amebae to erythrocytes and CHO cells, erythrop...

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“…3). The hgl probe hybridized with five to seven bands, depending on the strain, as has been previously reported (35).…”

Section: Vol 69 2001supporting

confidence: 78%

Section: Vol 69 2001mentioning

confidence: 99%

mentioning

confidence: 99%

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Intermediate Subunit of the Gal/GalNAc Lectin of Entamoeba histolytica Is a Member of a Gene Family Containing Multiple CXXC Sequence Motifs

et al. 2001

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“…The Gal/GalNAc lectin is a 260-kDa heterodimer consisting of heavy (170-kDa) and light (31-to 35-kDa) subunits linked by disulfide bonds (154,155) and noncovalently associated with a 150-kDa intermediate subunit (40). The heavy, intermediate, and light subunits are encoded by multiple gene families (168). The heavy subunit is encoded by a family of five genes, hgl1 to hgl5 (168), located at five district loci, while the light subunit (31/35 kDa) is encoded by a family of six or seven lgl genes in the genome (132,168,217).…”

Section: Pathogenicitymentioning

confidence: 99%

Laboratory Diagnosis of Amebiasis

2003

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The detection of Entamoeba histolytica, the causative agent of amebiasis, is an important goal of the clinical microbiology laboratory. To assess the scope of E. histolytica infection, it is necessary to utilize accurate diagnostic tools. As more is discovered about the molecular and cell biology of E. histolytica, there is great potential for further understanding the pathogenesis of amebiasis. Molecular biology-based diagnosis may become the technique of choice in the future because establishment of these protozoa in culture is still not a routine clinical laboratory process. In all cases, combination of serologic tests with detection of the parasite (by antigen detection or PCR) offers the best approach to diagnosis, while PCR techniques remain impractical in many developing country settings. The detection of amebic markers in serum in patients with amebic colitis and liver abscess appears promising but is still only a research tool. On the other hand, stool antigen detection tests offer a practical, sensitive, and specific way for the clinical laboratory to detect intestinal E. histolytica. All the current tests suffer from the fact that the antigens detected are denatured by fixation of the stool specimen, limiting testing to fresh or frozen samples

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“…The light subunit shows two 35-and 31-kDa isoforms, with the latter having a glycosylphosphatidylinositol anchor. This subunit is also encoded by a gene family that have 79 to 85% nucleotide sequence identity (123). Its function is not yet clear but seems to involve the modulation of parasite cytopathic activity (7,109).…”

Section: Early Invasive Lesions With Superficial Ulcerationmentioning

confidence: 99%