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1.
The protozoan parasite Trypanosoma cruzi, the agent of Chagas disease, has a large number of mucin molecules on its surface, whose expression is regulated during the life cycle. These mucins are the main acceptors of sialic acid, a monosaccharide that is required by the parasite to infect and survive in the mammalian host. A large mucin-like gene family named TcMUC containing about 500 members has been identified previously in T. cruzi. TcMUC can be divided into two subfamilies according to the presence or absence of tandem repeats in the central region of the genes. In this work, T. cruzi parasites were transfected with one tagged member of each subfamily. Only the product from the gene with repeats was highly O-glycosylated in vivo. The O-linked oligosaccharides consisted mainly of beta-d-Galp(1-->4)GlcNAc and beta-d-Galp(1-->4)[beta-d-Galp(1-->6)]-d-GlcNAc. The same glycosyl moieties were found in endogenous mucins. The mature product was anchored by glycosylphosphatidylinositol to the plasma membrane and exposed to the medium. Sera from infected mice recognized the recombinant product of one repeats-containing gene thus showing that they are expressed during the infection. TcMUC genes encode a hypervariable region at the N terminus. We now show that the hypervariable region is indeed present in the exposed mature N termini of the mucins because sera from infected hosts recognized peptides having sequences from this region. The results are discussed in comparison with the mucins from the insect stages of the parasite (Di Noia, J. M., D'Orso, I., Sánchez, D. O., and Frasch, A. C. C. (2000) J. Biol. Chem. 275, 10218-10227) which do not have variable regions.  相似文献   

2.
The surface of the protozoan parasite Trypanosoma cruzi is covered in mucins, which contribute to parasite protection and to the establishment of a persistent infection. Their importance is highlighted by the fact that the approximately 850 mucin-encoding genes comprise approximately 1% of the parasite genome and approximately 6% of all predicted T. cruzi genes. The coordinate expression of a large repertoire of mucins containing variable regions in the mammal-dwelling stages of the T. cruzi life cycle suggests a possible strategy to thwart the host immune response. Here, we discuss the expression profiling of T. cruzi mucins, the mechanisms leading to the acquisition of mucin diversity and the possible consequences of a mosaic surface coat in the interplay between parasite and host.  相似文献   

3.
Trypanosoma cruzi is wrapped by a dense coat of mucin-type molecules encoded by complex gene families termed TcSMUG and TcMUC, which are expressed in the insect- and mammal-dwelling forms of the parasite, respectively. Here, we dissect the contribution of distinct post-translational modifications on the trafficking of these glycoconjugates. In vivo tracing and characterization of tagged-variants expressed by transfected epimastigotes indicate that although the N-terminal signal peptide is responsible for targeting TcSMUG products to the endoplasmic reticulum (ER), the glycosyl phosphatidylinositol (GPI)-anchor likely functions as a forward transport signal for their timely progression along the secretory pathway. GPI-minus variants accumulate in the ER, with only a minor fraction being ultimately released to the medium as anchorless products. Secreted products, but not ER-accumulated ones, display several diagnostic features of mature mucin-type molecules including extensive O-type glycosylation, Galf-based epitopes recognized by monoclonal antibodies, and terminal Galp residues that become readily sialylated upon addition of parasite trans-sialidases. Processing of N-glycosylation site(s) is dispensable for the overall TcSMUG mucin-type maturation and secretion. Despite undergoing different O-glycosylation elaboration, TcMUC reporters yielded quite similar results, thus indicating that (i) molecular trafficking signals are structurally and functionally conserved between mucin families, and (ii) TcMUC and TcSMUG products are recognized and processed by a distinct repertoire of stage-specific glycosyltransferases. Thus, using the fidelity of a homologous expression system, we have defined some biosynthetic aspects of T. cruzi mucins, key molecules involved in parasite protection and virulence.  相似文献   

4.
The surface of the protozoan Trypanosoma cruzi is covered by a dense coat of mucin-type glycoconjugates, which make a pivotal contribution to parasite protection and host immune evasion. Their importance is further underscored by the presence of >1000 mucin-like genes in the parasite genome. In the present study we demonstrate that one such group of genes, termed TcSMUG L, codes for previously unrecognized mucin-type glycoconjugates anchored to and secreted from the surface of insect-dwelling epimastigotes. These features are supported by the in vivo tracing and characterization of endogenous TcSMUG L products and recombinant tagged molecules expressed by transfected parasites. Besides displaying substantial homology to TcSMUG S products, which provide the scaffold for the major Gp35/50 mucins also present in insect-dwelling stages of the T. cruzi lifecycle, TcSMUG L products display unique structural and functional features, including being completely refractory to sialylation by parasite trans-sialidases. Although quantitative real time-PCR and gene sequencing analyses indicate a high degree of genomic conservation across the T. cruzi species, TcSMUG L product expression and processing is quite variable among different parasite isolates.  相似文献   

5.
A dense glycocalix covers the surface of Trypanosoma cruzi, the agent of Chagas disease. Sialic acid in the surface of the parasite plays an important role in the infectious process, however, T. cruzi is unable to synthesize sialic acid or the usual donor CMP-sialic acid. Instead, T. cruzi expresses a unique enzyme, the trans-sialidase (TcTS) involved in the transfer of sialic acid from host glycoconjugates to mucins of the parasite. The mucins are the major glycoproteins in the insect stage epimastigotes and in the infective trypomastigotes. Both, the mucins and the TcTS are anchored to the plasma membrane by a glycosylphosphatidylinositol anchor. Thus, TcTS may be shed into the bloodstream of the mammal host by the action of a parasite phosphatidylinositol-phospholipase C, affecting the immune system. The composition and structure of the sugars in the parasite mucins is characteristic of each differentiation stage, also, interstrain variations were described for epimastigote mucins. This review focus on the characteristics of the interplay between the trans-sialidase and the mucins of T. cruzi and summarizes the known carbohydrate structures of the mucins.  相似文献   

6.
7.
The surface of Trypanosoma cruzi is covered by different groups of mucins that are differentially expressed during the parasite life cycle. We have previously identified the major mucins from the bloodstream trypomastigote stage. Here, we present additional evidence that together with our previous observations allows for the identification of a second mucin group also expressed in the mammal-dwelling stages, but predominant in the intracellular amastigote. These mucins are encoded by many genes, are mostly composed of tandem repeats and are highly conserved except for an exposed hypervariable (HV) N-terminal peptide. Antibodies against HV-peptides are restricted to approximately 50% of the chronically infected human population, are monospecific (i.e. directed towards a single HV), and display low-avidity. In contrast, immunization with a single HV-peptide triggers high-avidity, cross-reacting humoral responses against multiple HV sequences, but not against other T. cruzi surface antigens. The diversity present in the HV regions and the characteristics of the antibody response against them suggest a role of these molecules in eluding and/or modulating the mammalian host immune system.  相似文献   

8.
9.
The poly(A)-binding protein (PABP), bound to the 3' poly(A) tail of eukaryotic mRNAs, plays critical roles in mRNA translation and stability. PABP autoregulates its synthesis by binding to a conserved A-rich sequence present in the 5'-untranslated region of PABP mRNA and repressing its translation. PABP is composed of two parts: the highly conserved N terminus, containing 4 RNA recognition motifs (RRMs) responsible for poly(A) and eIF4G binding; and the more variable C terminus, which includes the recently described PABC domain, and promotes intermolecular interaction between PABP molecules as well as cooperative binding to poly(A). Here we show that, in vitro, GST-PABP represses the translation of reporter mRNAs containing 20 or more A residues in their 5'-untranslated regions and remains effective as a repressor when an A61 tract is placed at different distances from the cap, up to 126 nucleotides. Deletion of the PABP C terminus, but not the PABC domain alone, significantly reduces its ability to inhibit translation when bound to sequences distal to the cap, but not to proximal ones. Moreover, cooperative binding by multiple PABP molecules to poly(A) requires the C terminus, but not the PABC domain. Further analysis using pull-down assays shows that the interaction between PABP molecules, mediated by the C terminus, does not require the PABC domain and is enhanced by the presence of RRM 4. In vivo, fusion proteins containing parts of the PABP C terminus fused to the viral coat protein MS2 have an enhanced ability to prevent the expression of chloramphenicol acetyltransferase reporter mRNAs containing the MS2 binding site at distal distances from the cap. Altogether, our results identify a proline- and glutamine-rich linker located between the RRMs and the PABC domain as being strictly required for PABP/PABP interaction, cooperative binding to poly(A) and enhanced translational repression of reporter mRNAs in vitro and in vivo.  相似文献   

10.
Leishmania parasites secrete a variety of proteins that are modified by phosphoglycan chains structurally similar to those of the cell surface glycolipid lipophosphoglycan. These proteins are collectively called proteophosphoglycans. We report here the cloning and sequencing of a novel Leishmania major proteophosphoglycan gene, ppg1. It encodes a large polypeptide of approximately 2300 amino acids. The N-terminal domain of approximately 70 kDa exhibits 11 imperfect amino acid repeats that show some homology to promastigote surface glycoproteins of the psa2/gp46 complex. The large central domain apparently consists exclusively of approximately 100 repetitive peptides of the sequence APSASSSSA(P/S)SSSSS(+/-S). Gene fusion experiments demonstrate that these peptide repeats are the targets of phosphoglycosylation in Leishmania and that they form extended filamentous structures reminiscent of mammalian mucins. The C-terminal domain contains a functional glycosylphosphatidylinositol anchor addition signal sequence, which confers cell surface localization to a normally secreted Leishmania acid phosphatase, when fused to its C terminus. Antibody binding studies show that the ppg1 gene product is phosphoglycosylated by phosphoglycan repeats and cap oligosaccharides. In contrast to previously characterized proteophosphoglycans, the ppg1 gene product is predominantly membrane-associated and it is expressed on the promastigote cell surface. Therefore this membrane-bound proteophosphoglycan may be important for direct host-parasite interactions.  相似文献   

11.
Parasite-derived mucin-like molecules might be involved in parasite attachment to and invasion of host cells. In addition, parasites might secrete mucin-degrading enzymes, enabling the penetration of protective mucus gels that overlie the mucosal surfaces of their potential hosts. Furthermore, they might generate binding ligands on the membrane-bound mucins of host cells by using specific glycosidases. It is possible that host mucins and mucin-like molecules prevent the establishment of parasites or facilitate parasite expulsion. They might also serve as a source of metabolic energy and adhesion ligands for those parasites adapted to exploit them. Sally Hicks and colleagues here review the biochemical properties of mucins and mucin-like molecules in relation to interactions (established and putative) between protozoan parasites and their hosts.  相似文献   

12.
Dsl1p is required for Golgi-endoplasmic reticulum (ER) retrograde transport in yeast. It interacts with the ER resident protein Tip20p and with delta-COP, a subunit of coatomer, the coat complex of COPI vesicles. To test the significance of these interactions, we mapped the different binding sites and created mutant versions of Dsl1p and delta-COP, which are unable to bind directly to each other. Three domains were identified in Dsl1p: a Tip20p binding region within the N-terminal 200 residues, a highly acidic region in the center of Dsl1p containing crucial tryptophan residues that is required for binding to delta-COP and essential for viability, and an evolutionarily well conserved domain at the C terminus. Most importantly, Dsl1p uses the same central acidic domain to interact not only with delta-COP but also with alpha-COP. Strong interaction with alpha-COP requires the presence of comparable amounts of epsilon-COP or beta' -COP. Thus, the binding characteristics of Dsl1p resemble those of many accessory factors of the clathrin coat. They interact with different layers of the vesicle coat by using tandemly arranged sequence motifs, some of which have dual specificity.  相似文献   

13.
Some parasites express mucin-like molecules. These have possible roles in attachment and invasion of host cells and in the avoidance of host immune processes. Enzymes of parasite origin might also facilitate infection, either by degrading host mucus barriers or by generating binding sites on host cells. Host mucins have roles in preventing parasite establishment or in parasite expulsion. They, in turn, might be exploited by parasites, either as sources of fuel or binding sites, or as host-finding targets. Here, we describe the biochemical properties of mucins and mucin-like molecules in relation to interactions (established and putative) between helminth parasites and their hosts.  相似文献   

14.
15.
16.
Formation of COPII-coated vesicles at the endoplasmic reticulum (ER) requires assembly onto the membrane of five cytosolic coat proteins, Sec23p, Sec24p, Sec13p, Sec31p, and Sar1p. A sixth vesicle coat component, Sec16p, is tightly associated with the ER membrane and has been proposed to act as a scaffold for membrane association of the soluble coat proteins. We previously showed that Sec23p binds to the C-terminal region of Sec16p. Here we use two-hybrid and coprecipitation assays to demonstrate that the essential COPII protein Sec24p binds to the central region of Sec16p. In vitro reconstitution of binding with purified recombinant proteins demonstrates that the interaction of Sec24p with the central domain of Sec16p does not depend on the presence of Sec23p. However, Sec23p facilitates binding of Sec24p to Sec16p, and the three proteins can form a ternary complex in vitro. Truncations of Sec24p demonstrate that the N-terminal and C-terminal regions of Sec24p display different binding specificities. The C terminus binds to the central domain of Sec16p, whereas the N terminus of Sec24p binds to both the central domain of Sec16p and to Sec23p. These findings define binding to Sec16p as a new function for Sec24p and support the idea that Sec16p organizes assembly of the COPII coat.  相似文献   

17.
18.
Secreted epithelial mucins are large macromolecules which exhibit extreme polydispersity, the molecular basis of which is not fully understood. We have obtained partial sequences of two genes (BSM1 and BSM2) coding for two distinct molecules. This is the first time that such closely-related genes have been identified for any mucin from an animal. We propose that a combination of multiple homologous genes, alternative splicing, differential glycosylation, and additional post-translational processing all contribute to the extreme polydispersity of mucins. The multiple domain structure and non-identical tandem repeats are also very important for the generation of the saccharide diversities of mucins.  相似文献   

19.
The invasion of host cells by the malaria parasite Plasmodium falciparum requires specific protein-protein interactions between parasite and host receptors and an intracellular translocation machinery to power the process. The transmembrane erythrocyte binding protein-175 (EBA-175) and thrombospondin-related anonymous protein (TRAP) play central roles in this process. EBA-175 binds to glycophorin A on human erythrocytes during the invasion process, linking the parasite to the surface of the host cell. In this report, we show that the cytoplasmic domain of EBA-175 encodes crucial information for its role in merozoite invasion, and that trafficking of this protein is independent of this domain. Further, we show that the cytoplasmic domain of TRAP, a protein that is not expressed in merozoites but is essential for invasion of liver cells by the sporozoite stage, can substitute for the cytoplasmic domain of EBA-175. These results show that the parasite uses the same components of its cellular machinery for invasion regardless of the host cell type and invasive form.  相似文献   

20.
MUC17, a novel membrane-tethered mucin   总被引:13,自引:0,他引:13  
Membrane mucins have several functions in epithelial cells including cytoprotection, extravasation during metastases, maintenance of luminal structure, and signal transduction. In this paper we describe a large membrane mucin expressed in the normal intestine. This novel mucin, designated MUC17, contains an extended, repetitive extracellular glycosylation domain and a carboxyl terminus with two EGF-like domains, a SEA module domain, a transmembrane domain, and a cytoplasmic domain with potential serine and tyrosine phosphorylation sites. RNA blot analysis and in situ hybridization indicates that MUC17 is expressed in select pancreatic and colon cancer cell lines and in intestinal absorptive cells. Radiation hybrid mapping localized MUC17 to chromosome 7q22 where it resides in close proximity with three other membrane mucin genes, MUC3A, MUC3B, and MUC12. Thus, these membrane mucins reside together in a gene cluster, but are expressed in different tissues and are likely to have different functions as well.  相似文献   

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