scholarly journals Trichomonas vaginalis Polyamine Metabolism Is Linked to Host Cell Adherence and Cytotoxicity

2005 ◽  
Vol 73 (5) ◽  
pp. 2602-2610 ◽  
Author(s):  
Ana F. Garcia ◽  
M. Benchimol ◽  
J. F. Alderete
Keyword(s):  
Host Cell ◽  
Trichomonas Vaginalis ◽  
Cysteine Proteinase ◽  
Growth Arrest ◽  
Electron Microscopic Examination ◽  
Host Cells ◽  
Polyamine Metabolism ◽  
Cell Adherence ◽  
Electron Microscopic ◽  
Control Versus

ABSTRACT Trichomonas vaginalis secretes putrescine that is readily detected in vaginal secretions. We wanted to examine the effect of decreased putrescine synthesis by inhibition of ornithine decarboxylase (ODC) on T. vaginalis. One reason is because inhibition of Tritrichomonas foetus ODC results in growth arrest, destruction of hydrogenosomes, and decreased amounts of hydrogenosomal enzymes. Treatment of T. vaginalis T016 with ≥20 mM 1,4-diamino-2-butanone (DAB) to inhibit ODC resulted in growth arrest, which was reversed by addition of exogenous putrescine. No similar reversal of growth arrest was achieved with the polyamines spermine or spermidine or with iron. Electron microscopic examination of control versus DAB-treated trichomonads did not reveal any adverse effects on the number and integrity of hydrogenosomes. Further, the adhesins AP65, AP51, and AP33 mediating binding to immortalized vaginal epithelial cells (VECs) share identity to enzymes of the hydrogenosome organelle, and there was no difference in amounts of adhesins between control versus DAB-treated T. vaginalis parasites. Likewise, similar patterns and extent of fluorescence were evident for the prominent AP65 adhesin. Surprisingly, DAB treatment increased by 4- to 20-fold above untreated trichomonads handled identically the level of adherence mediated by adhesins. Interestingly, the enhanced attachment to VECs was reversed by exogenous putrescine added to DAB-treated trichomonads. Equally noteworthy was that DAB-treated T. vaginalis with enhanced adherence did not possess the previously reported ability to kill host cells in a contact-dependent fashion mediated by cysteine proteinases, and total cysteine proteinase activity patterns were identical between control and DAB-treated trichomonads. Overall, these data suggest that polyamine metabolism and secreted putrescine are linked to host cell adherence and cytotoxicity.

scholarly journals A Novel Trichomonas vaginalis Surface Protein Modulates Parasite Attachment via Protein:Host Cell Proteoglycan Interaction

mBio ◽  
2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Brenda M. Molgora ◽  
Anand Kumar Rai ◽  
Michael J. Sweredoski ◽  
Annie Moradian ◽  
Sonja Hess ◽  
...  
Keyword(s):  
Host Cell ◽  
Trichomonas Vaginalis ◽  
Surface Protein ◽  
Host Cells ◽  
Parasite Line ◽  
Cell Adherence ◽  
Common Component ◽  
Content Type ◽  
Sexually Transmitted ◽  
Significant Difference

ABSTRACT Trichomonas vaginalis is a highly prevalent, sexually transmitted parasite which adheres to mucosal epithelial cells to colonize the human urogenital tract. Despite adherence being crucial for this extracellular parasite to thrive within the host, relatively little is known about the mechanisms or key molecules involved in this process. Here, we have identified and characterized a T. vaginalis hypothetical protein, TVAG_157210 (TvAD1), as a surface protein that plays an integral role in parasite adherence to the host. Quantitative proteomics revealed TvAD1 to be ∼4-fold more abundant in parasites selected for increased adherence (MA parasites) than the isogenic parental (P) parasite line. De novo modeling suggested that TvAD1 binds N-acetylglucosamine (GlcNAc), a sugar comprising host glycosaminoglycans (GAGs). Adherence assays utilizing GAG-deficient cell lines determined that host GAGs, primarily heparan sulfate (HS), mediate adherence of MA parasites to host cells. TvAD1 knockout (KO) parasites, generated using CRISPR-Cas9, were found to be significantly reduced in host cell adherence, a phenotype that is rescued by overexpression of TvAD1 in KO parasites. In contrast, there was no significant difference in parasite adherence to GAG-deficient lines by KO parasites compared with wild-type, which is contrary to that observed for KO parasites overexpressing TvAD1. Isothermal titration calorimetric (ITC) analysis showed that TvAD1 binds to HS, indicating that TvAD1 mediates host cell adherence via HS interaction. In addition to characterizing the role of TvAD1 in parasite adherence, these studies reveal a role for host GAG molecules in T. vaginalis adherence. IMPORTANCE The ability of the sexually transmitted parasite Trichomonas vaginalis to adhere to its human host is critical for establishing and maintaining an infection. Yet how parasites adhere to host cells is poorly understood. In this study, we employed a novel adherence selection method to identify proteins involved in parasite adherence to the host. This method led to the identification of a protein, with no previously known function, that is more abundant in parasites with increased capacity to bind host cells. Bioinformatic modeling and biochemical analyses revealed that this protein binds a common component on the host cell surface proteoglycans. Subsequent creation of parasites that lack this protein directly demonstrated that the protein mediates parasite adherence via an interaction with host cell proteoglycans. These findings both demonstrate a role for this protein in T. vaginalis adherence to the host and shed light on host cell molecules that participate in parasite colonization.

scholarly journals A Novel Cadherin-like Protein Mediates Adherence to and Killing of Host Cells by the Parasite Trichomonas vaginalis

mBio ◽  
2019 ◽  
Vol 10 (3) ◽  
Author(s):  
Yi-Pei Chen ◽  
Angelica M. Riestra ◽  
Anand Kumar Rai ◽  
Patricia J. Johnson
Keyword(s):  
Cell Adhesion ◽  
Binding Site ◽  
Host Cell ◽  
Calcium Binding ◽  
Trichomonas Vaginalis ◽  
Host Cells ◽  
Wild Type ◽  
Cell Adherence ◽  
Calcium Binding Site ◽  
Content Type

ABSTRACT Trichomonas vaginalis, a prevalent sexually transmitted parasite, adheres to and induces cytolysis of human mucosal epithelial cells. We have characterized a hypothetical protein, TVAG_393390, with predicted tertiary structure similar to that of mammalian cadherin proteins involved in cell-cell adherence. TVAG_393390, renamed cadherin-like protein (CLP), contains a calcium-binding site at a position conserved in cadherins. CLP is surface localized, and its mRNA and protein levels are significantly upregulated upon parasite adherence to host cells. To test the roles of CLP and its calcium-binding dependency during host cell adherence, we first demonstrated that wild-type CLP (CLP) binds calcium with a high affinity, whereas the calcium-binding site mutant protein (CLP-mut) does not. CLP and CLP-mut constructs were then used to overexpress these proteins in T. vaginalis. Parasites overexpressing CLP have ∼3.5-fold greater adherence to host cells than wild-type parasites, and this increased adherence is ablated by mutating the calcium-binding site. Additionally, competition with recombinant CLP decreased parasite binding to host cells. We also found that overexpression of CLP induced parasite aggregation which was further enhanced in the presence of calcium, whereas CLP-mut overexpression did not affect aggregation. Lastly, parasites overexpressing wild-type CLP induced killing of host cells ∼2.35-fold, whereas parasites overexpressing CLP-mut did not have this effect. These analyses describe the first parasitic CLP and demonstrate a role for this protein in mediating parasite-parasite and host-parasite interactions. T. vaginalis CLP may represent convergent evolution of a parasite protein that is functionally similar to the mammalian cell adhesion protein cadherin, which contributes to parasite pathogenesis. IMPORTANCE The adherence of pathogens to host cells is critical for colonization of the host and establishing infection. Here we identify a protein with no known function that is more abundant on the surface of parasites that are better at binding host cells. To interrogate a predicted function of this protein, we utilized bioinformatic protein prediction programs which allowed us to uncover the first cadherin-like protein (CLP) found in a parasite. Cadherin proteins are conserved metazoan proteins with central roles in cell-cell adhesion, development, and tissue structure maintenance. Functional characterization of this CLP from the unicellular parasite Trichomonas vaginalis demonstrated that the protein mediates both parasite-parasite and parasite-host adherence, which leads to an enhanced killing of host cells by T. vaginalis. Our findings demonstrate the presence of CLPs in unicellular pathogens and identify a new host cell binding protein family in a human-infective parasite.

Cell interactions of Listeria monocytogenes L forms and peritoneal exudative cells in rats

1993 ◽  
Vol 39 (11) ◽  
pp. 1014-1021 ◽  
Author(s):  
L. Mihailova ◽  
N. Markova ◽  
T. Radoucheva ◽  
D. Veljanov ◽  
S. Radoevska
Keyword(s):  
Listeria Monocytogenes ◽  
Peritoneal Cavity ◽  
Electron Microscopic Examination ◽  
Lavage Fluid ◽  
Host Cells ◽  
Electron Microscopic ◽  
Cellular Defense ◽  
Scanning Electron Microscopic ◽  
Scanning Electron

Listeria monocytogenes 4b and its forms without cell walls (L forms of a protoplastic type) were used to study in vivo interactions with host cells. Samples of peritoneal lavage fluid were obtained from rats intraperitoneally inoculated at intervals between 1 and 15 days after challenge, for scanning electron microscopic, bacteriological, biochemical, and cytometrical investigations. Scanning electron microscopic examination revealed continuous adhesion of L forms on the macrophage surface up to 15 days after inoculation. The persistence of the L forms within the peritoneal cavity was also shown bacteriologically at all sample times, while the parental bacterial forms were isolated from the peritoneal cavity up to 7 days after challenge. The total count of peritoneal exudative cells determined by automated flow peroxidase cytometry peaked on the 15th day in animals infected with parental forms, while in animals infected with L forms the peak was lower and the macrophage population was predominant. The glycolytic and acid phosphatase activity of peritoneal exudative cells was two times higher in rats infected with L forms as compared with rats infected with the L. monocytogenes parental forms on the 3rd day after challenge. An understanding of the nature of the interactions between L forms of L. monocytogenes and peritoneal exudative cells found in vivo could be used to establish the influence of L forms on host cellular defense mechanisms.Key words: Listeria monocytogenes, L forms, peritoneal exudative cells, electron microscopy.

Structure and host–actinomycete interactions in developing root nodules of Comptonia peregrina

1978 ◽  
Vol 56 (5) ◽  
pp. 502-531 ◽  
Author(s):  
William Newcomb ◽  
R. L. Peterson ◽  
Dale Callaham ◽  
John G. Torrey
Keyword(s):  
Host Cell ◽  
Root Nodules ◽  
Host Cells ◽  
Electron Microscopic ◽  
Cortical Cells ◽  
Host Cytoplasm ◽  
Transmission Electron ◽  
Microscopic Studies ◽  
Host Plasma Membrane ◽  
Soil Actinomycete

Correlated fluorescence, bright-field, transmission electron, and scanning electron microscopic studies were made on developing root nodules of Comptonia peregrina (L.) Coult. (Myricaceae) produced by a soil actinomycete which invades the root and establishes a symbiosis leading to fixation of atmospheric dinitrogen. After entering the host via a root hair infection, the hyphae of the endophyte perforate root cortical cells by local degradation of host cell walls and penetration of the host cytoplasm. The intracellular hyphae are always surrounded by host plasma membrane and a thick polysaccharide material termed the capsule. (For convenience, term intracellular refers to the endophyte being inside a Comptonia cell as distinguished from being intercellular, i.e.. between host cells, even though the former is actually extracellular as the endophyte is separated from the host cytoplasm by the host plasmalemma.) Numerous profiles of vesiculate rough endoplasmic reticulum (RER) occur near the growing hyphae. Although the capsule shows a positive Thiery reaction indicating its polysaccharide nature, the fibrillar contents of the RER do not, leaving uncertain whether the capsule results from polymers derived from the RER. Amyloplasts of the cortical cells lose their starch deposits during hyphal proliferation. The hyphae branch extensively in specific layers of the cortex, penetrating much of the host cytoplasm. At this stage, hyphal ends become swollen and form septate club-shaped vesicles within the periphery of the host cells. Lipid-like inclusions and Thiery-positive particles, possibly glycogen, are observed in the hyphae at this time. Associated with hyphal development is an increase in average host cell volume, although nuclear volume appears to remain constant. Concomitant with vesicle maturation, the mitochondrial population increases sharply, suggesting a possible relationship to vesicle function. The intimate interactions between host and endophyte during development of the symbiotic relationship are emphasized throughout.

scholarly journals Aziridine-2,3-Dicarboxylate-Based Cysteine Cathepsin Inhibitors Induce Cell Death in Leishmania major Associated with Accumulation of Debris in Autophagy-Related Lysosome-Like Vacuoles

2010 ◽  
Vol 54 (12) ◽  
pp. 5028-5041 ◽  
Author(s):  
Uta Schurigt ◽  
Caroline Schad ◽  
Christin Glowa ◽  
Ulrike Baum ◽  
Katja Thomale ◽  
...  
Keyword(s):  
Cell Death ◽  
Leishmania Major ◽  
Cysteine Proteinase ◽  
Membrane Integrity ◽  
New Drugs ◽  
Host Cells ◽  
Electron Microscopic ◽  
Cysteine Cathepsins ◽  
Cysteine Cathepsin ◽  
Microscopic Studies

ABSTRACT The papain-like cysteine cathepsins expressed by Leishmania play a key role in the life cycle of these parasites, turning them into attractive targets for the development of new drugs. We previously demonstrated that two compounds of a series of peptidomimetic aziridine-2,3-dicarboxylate [Azi(OBn)2]-based inhibitors, Boc-(S)-Leu-(R)-Pro-(S,S)-Azi(OBn)2 (compound 13b) and Boc-(R)-Leu-(S)-Pro-(S,S)-Azi(OBn)2 (compound 13e), reduced the growth and viability of Leishmania major and the infection rate of macrophages while not showing cytotoxicity against host cells. In the present study, we characterized the mode of action of inhibitors 13b and 13e in L. major. Both compounds targeted leishmanial cathepsin B-like cysteine cathepsin cysteine proteinase C, as shown by fluorescence proteinase activity assays and active-site labeling with biotin-tagged inhibitors. Furthermore, compounds 13b and 13e were potent inducers of cell death in promastigotes, characterized by cell shrinkage, reduction of mitochondrial transmembrane potential, and increased DNA fragmentation. Transmission electron microscopic studies revealed the enrichment of undigested debris in lysosome-like organelles participating in micro- and macroautophagy-like processes. The release of digestive enzymes into the cytoplasm after rupture of membranes of lysosome-like vacuoles resulted in the significant digestion of intracellular compartments. However, the plasma membrane integrity of compound-treated promastigotes was maintained for several hours. Taken together, our results suggest that the induction of cell death in Leishmania by cysteine cathepsin inhibitors 13b and 13e is different from mammalian apoptosis and is caused by incomplete digestion in autophagy-related lysosome-like vacuoles.

scholarly journals Intracellular Staphylococcus aureusEscapes the Endosome and Induces Apoptosis in Epithelial Cells

1998 ◽  
Vol 66 (1) ◽  
pp. 336-342 ◽  
Author(s):  
Kenneth W. Bayles ◽  
Carla A. Wesson ◽  
Linda E. Liou ◽  
Lawrence K. Fox ◽  
Gregory A. Bohach ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Nuclear Dna ◽  
Electron Microscopic Examination ◽  
Intracellular Survival ◽  
Host Cells ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Epithelial Cell Line ◽  
Electron Microscopic ◽  
Endosomal Membrane

ABSTRACT We examined the invasion of an established bovine mammary epithelial cell line (MAC-T) by a Staphylococcus aureusmastitis isolate to study the potential role of intracellular survival in the persistence of staphylococcal infections. S. aureuscells displayed dose-dependent invasion of MAC-T cells and intracellular survival. An electron microscopic examination of infected cells indicated that the bacteria induced internalization via a mechanism involving membrane pseudopod formation and then escaped into the cytoplasm following lysis of the endosomal membrane. Two hours after the internalization of S. aureus, MAC-T cells exhibited detachment from the matrix, rounding, a mottled cell membrane, and vacuolization of the cytoplasm, all of which are indicative of cells undergoing programmed cell death (apoptosis). By 18 h, the majority of the MAC-T cell population exhibited an apoptotic morphology. Other evidence for apoptosis was the generation of MAC-T cell DNA fragments differing in size by increments of approximately 180 bp and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling of the fragmented nuclear DNA of the infected host cells. These results demonstrate that after internalizationS. aureus escapes the endosome and induces apoptosis in nonprofessional phagocytes.

scholarly journals African Swine Fever Virus Proteinase Is Essential for Core Maturation and Infectivity

2003 ◽  
Vol 77 (10) ◽  
pp. 5571-5577 ◽  
Author(s):  
Alí Alejo ◽  
Germán Andrés ◽  
María L. Salas
Keyword(s):  
African Swine Fever Virus ◽  
Sequence Similarity ◽  
Cysteine Proteinase ◽  
African Swine Fever ◽  
Virus Production ◽  
Electron Microscopic Examination ◽  
Proteolytic Processing ◽  
Fever Virus ◽  
Core Shell ◽  
Electron Microscopic

ABSTRACT African swine fever virus (ASFV) encodes two polyprotein precursors named pp220 and pp62 that are sequentially processed during viral infection, giving rise to six major structural proteins. These reside at the core shell, a matrix domain located between the endoplasmic reticulum-derived inner envelope and the DNA-containing nucleoid. Proteolytic processing of the polyprotein precursors is catalyzed by the viral proteinase pS273R, a cysteine proteinase that shares sequence similarity with the SUMO1-processing peptidases. We describe here the construction and characterization of an ASFV recombinant, vS273Ri, that inducibly expresses the ASFV proteinase. Using vS273Ri, we show that repression of proteinase expression inhibits polyprotein processing and strongly impairs infective virus production. Electron microscopic examination of vS273Ri-infected cells showed that inhibition of proteolytic processing leads to the assembly of defective icosahedral particles containing a noncentered electron-dense nucleoid surrounded by an abnormal core shell of irregular thickness. The analysis of purified extracellular defective particles revealed that they contain the unprocessed pp220 and pp62 precursors, as well as the major DNA-binding nucleoid proteins p10 and pA104R. Altogether, these results indicate that the proteolytic processing of the polyproteins is not required for their incorporation into the assembling particles nor for the incorporation of the DNA-containing nucleoid. Instead, the ASFV proteinase is involved in a late maturational step that is essential for proper core assembly and infectivity.

scholarly journals Bartonella quintana Variably Expressed Outer Membrane Proteins Mediate Vascular Endothelial Growth Factor Secretion but Not Host Cell Adherence

2006 ◽  
Vol 74 (9) ◽  
pp. 5003-5013 ◽  
Author(s):  
Berit Schulte ◽  
Dirk Linke ◽  
Sandra Klumpp ◽  
Martin Schaller ◽  
Tanja Riess ◽  
...  
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Growth Factor ◽  
Host Cell ◽  
Outer Membrane ◽  
Outer Membrane Proteins ◽  
Host Cells ◽  
Vascular Endothelial ◽  
Cell Adherence ◽  
Bartonella Quintana ◽  
Endothelial Growth Factor

ABSTRACT Bartonella quintana causes trench fever, endocarditis, and the vasculoproliferative disorders bacillary angiomatosis and peliosis hepatis in humans. Little is known about the interaction of this pathogen with host cells. We attempted to elucidate the interaction of B. quintana with human macrophages (THP-1) and epithelial cells (HeLa 229). Remarkably, only B. quintana strain JK-31 induced secretion of vascular endothelial growth factor (VEGF) from THP-1 and HeLa 229 cells upon infection similar to the secretion induced by B. henselae Marseille, whereas other strains (B. quintana 2-D70, B. quintana Toulouse, and B. quintana Munich) did not induce such secretion. Immunofluorescence testing and electron microscopy revealed that the B. quintana strains unable to induce VEGF secretion did not express the variable outer membrane proteins (Vomps) on their surfaces. Surprisingly, the increase in VEGF secretion mediated by B. quintana JK-31 was not paralleled by elevated host cell adherence rates compared with the rates for Vomp-negative B. quintana strains. Our results suggest that the Vomps play a leading role in the angiogenic reprogramming of host cells by B. quintana but not in the adherence to host cells.

Ultrastructure of the haustorium of the peanut late leaf spot fungus Cercosporidium personatum

1989 ◽  
Vol 67 (4) ◽  
pp. 1198-1202 ◽  
Author(s):  
C. W. Mims ◽  
E. S. Luttrell ◽  
S. C. Alderman
Keyword(s):  
Host Cell ◽  
Host Cells ◽  
Electron Microscopic ◽  
Cercosporidium Personatum ◽  
Transmission Electron ◽  
Transmission Electron Microscopic ◽  
Biotrophic Fungi ◽  
Leaf Spot Fungus ◽  
Extrahaustorial Matrix ◽  
Cell Protoplast

Data from scanning and transmission electron microscopic observations support light microscopic reports of the production of haustoria by the hemibiotrophic fungus Cercosporidium personatum. The trunklike base of the haustorium extended a short distance into the host cell where it formed three to five slightly thinner primary branches. These branches terminated in multiple, smaller, mostly opposite branch tips that gave the end of the haustorium a coralloid appearance. The morphology of this haustorium was distinctly different from the more extensively studied haustoria of various biotrophic fungi. Haustoria of C. personatum were observed in both living and dead host cells. In living cells an extrahaustorial matrix and extrahaustorial membrane separated the haustorium wall from the host cell protoplast. In dead cells the extrahaustorial membrane was absent. Haustoria in dead cells remained intact and appeared healthy.

scholarly journals Trichomonas vaginalis extracellular vesicles are internalized by host cells using proteoglycans and caveolin-dependent endocytosis

2019 ◽  
Vol 116 (43) ◽  
pp. 21354-21360 ◽  
Author(s):  
Anand Kumar Rai ◽  
Patricia J. Johnson
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Extracellular Vesicles ◽  
Mammalian Cells ◽  
Trichomonas Vaginalis ◽  
Host Cells ◽  
Titration Calorimetry ◽  
Mammalian Host ◽  
Target Cells ◽  
Host Cell Surface

Trichomonas vaginalis, a human-infective parasite, causes the most prevalent nonviral sexually transmitted infection worldwide. This pathogen secretes extracellular vesicles (EVs) that mediate its interaction with host cells. Here, we have developed assays to study the interface between parasite EVs and mammalian host cells and to quantify EV internalization by mammalian cells. We show that T. vaginalis EVs interact with glycosaminoglycans on the surface of host cells and specifically bind to heparan sulfate (HS) present on host cell surface proteoglycans. Moreover, competition assays using HS or removal of HS from the host cell surface strongly inhibit EV uptake, directly demonstrating that HS proteoglycans facilitate EV internalization. We identified an abundant protein on the surface of T. vaginalis EVs, 4-α-glucanotransferase (Tv4AGT), and show using isothermal titration calorimetry that this protein binds HS. Tv4AGT also competitively inhibits EV uptake, defining it as an EV ligand critical for EV internalization. Finally, we demonstrate that T. vaginalis EV uptake is dependent on host cell cholesterol and caveolin-1 and that internalization proceeds via clathrin-independent, lipid raft-mediated endocytosis. These studies reveal mechanisms used to drive host:pathogen interactions and further our understanding of how EVs are internalized by target cells to allow cross-talk between different cell types.

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