<i>O</i> -fucosylated glycoproteins form assemblies in close proximity to the nuclear pore complexes of <i>Toxoplasma gondii</i>

Proceedings of the National Academy of Sciences · 2016-09-23 · 48 citations

Toxoplasma gondii is an intracellular parasite that causes disseminated infections in fetuses and immunocompromised individuals. Although gene regulation is important for parasite differentiation and pathogenesis, little is known about protein organization in the nucleus. Here we show that the fucose-binding Aleuria aurantia lectin (AAL) binds to numerous punctate structures in the nuclei of tachyzoites, bradyzoites, and sporozoites but not oocysts. AAL also binds to Hammondia and Neospora nuclei but not to more distantly related apicomplexans. Analyses of the AAL-enriched fraction indicate that AAL binds O-linked fucose added to Ser/Thr residues present in or adjacent to Ser-rich domains (SRDs). Sixty-nine Ser-rich proteins were reproducibly enriched with AAL, including nucleoporins, mRNA-processing enzymes, and cell-signaling proteins. Two endogenous SRDs-containing proteins and an SRD-YFP fusion localize with AAL to the nuclear membrane. Superresolution microscopy showed that the majority of the AAL signal localizes in proximity to nuclear pore complexes. Host cells modify secreted proteins with O-fucose; here we describe the O-fucosylation pathway in the nucleocytosol of a eukaryote. Furthermore, these results suggest O-fucosylation is a mechanism by which proteins involved in gene expression accumulate near the NPC.

Anti-Retroviral Lectins Have Modest Effects on Adherence of Trichomonas vaginalis to Epithelial Cells In Vitro and on Recovery of Tritrichomonas foetus in a Mouse Vaginal Model

PLoS ONE · 2015-08-07 · 29 citations

Trichomonas vaginalis causes vaginitis and increases the risk of HIV transmission by heterosexual sex, while Tritrichomonas foetus causes premature abortion in cattle. Our goals were to determine the effects, if any, of anti-retroviral lectins, which are designed to prevent heterosexual transmission of HIV, on adherence of Trichomonas to ectocervical cells and on Tritrichomonas infections in a mouse model. We show that Trichomonas Asn-linked glycans (N-glycans), like those of HIV, bind the mannose-binding lectin (MBL) that is part of the innate immune system. N-glycans of Trichomonas and Tritrichomonas bind anti-retroviral lectins (cyanovirin-N and griffithsin) and the 2G12 monoclonal antibody, each of which binds HIV N-glycans. Binding of cyanovirin-N appears to be independent of susceptibility to metronidazole, the major drug used to treat Trichomonas. Anti-retroviral lectins, MBL, and galectin-1 cause Trichomonas to self-aggregate and precipitate. The anti-retroviral lectins also increase adherence of ricin-resistant mutants, which are less adherent than parent cells, to ectocervical cell monolayers and to organotypic EpiVaginal tissue cells. Topical application of either anti-retroviral lectins or yeast N-glycans decreases by 40 to 70% the recovery of Tritrichomonas from the mouse vagina. These results, which are explained by a few simple models, suggest that the anti-retroviral lectins have a modest potential for preventing or treating human infections with Trichomonas.

Effects of N-glycan precursor length diversity on quality control of protein folding and on protein glycosylation

Seminars in Cell and Developmental Biology · 2014-12-02 · 59 citations

The glycosylation-dependent interaction of perlecan core protein with LDL: implications for atherosclerosis

Journal of Lipid Research · 2014-12-21 · 30 citations

Perlecan is a major heparan sulfate (HS) proteoglycan in the arterial wall. Previous studies have linked it to atherosclerosis. Perlecan contains a core protein and three HS side chains. Its core protein has five domains (DI–DV) with disparate structures and DII is highly homologous to the ligand-binding portion of LDL receptor (LDLR). The functional significance of this domain has been unknown. Here, we show that perlecan DII interacts with LDL. Importantly, the interaction largely relies on O-linked glycans that are only present in the secreted DII. Among the five repeat units of DII, most of the glycosylation sites are from the second unit, which is highly divergent and rich in serine and threonine, but has no cysteine residues. Interestingly, most of the glycans are capped by the negatively charged sialic acids, which are critical for LDL binding. We further demonstrate an additive effect of HS and DII on LDL binding. Unlike LDLR, which directs LDL uptake through endocytosis, this study uncovers a novel feature of the perlecan LDLR-like DII in receptor-mediated lipoprotein retention, which depends on its glycosylation. Thus, perlecan glycosylation may play a role in the early LDL retention during the development of atherosclerosis. Perlecan is a major heparan sulfate (HS) proteoglycan in the arterial wall. Previous studies have linked it to atherosclerosis. Perlecan contains a core protein and three HS side chains. Its core protein has five domains (DI–DV) with disparate structures and DII is highly homologous to the ligand-binding portion of LDL receptor (LDLR). The functional significance of this domain has been unknown. Here, we show that perlecan DII interacts with LDL. Importantly, the interaction largely relies on O-linked glycans that are only present in the secreted DII. Among the five repeat units of DII, most of the glycosylation sites are from the second unit, which is highly divergent and rich in serine and threonine, but has no cysteine residues. Interestingly, most of the glycans are capped by the negatively charged sialic acids, which are critical for LDL binding. We further demonstrate an additive effect of HS and DII on LDL binding. Unlike LDLR, which directs LDL uptake through endocytosis, this study uncovers a novel feature of the perlecan LDLR-like DII in receptor-mediated lipoprotein retention, which depends on its glycosylation. Thus, perlecan glycosylation may play a role in the early LDL retention during the development of atherosclerosis. CVD is, and will continue to be in the foreseeable future, the most common cause of death worldwide. In the United States, it kills more than 800,000 people annually (nearly one of every three deaths); the mortality is greater than any other disease (1Go A.S. Mozaffarian D. Roger V.L. Benjamin E.J. Berry J.D. Blaha M.J. Dai S. Ford E.S. Fox C.S. Franco S. et al.Heart disease and stroke statistics–2014 update: a report from the American Heart Association.Circulation. 2014; 129: e28-e292Crossref PubMed Scopus (4481) Google Scholar). The leading cause of CVD is atherosclerosis, which is a pathological condition arising from fibrous plaque build-up inside the arterial wall. The plaque narrows the lumen of blood vessels and restricts blood flow. Rupture of the advanced plaque induces the formation of thrombus and blocks blood flow, which results in complications such as heart attack or stroke (2Libby P. Ridker P.M. Hansson G.K. Progress and challenges in translating the biology of atherosclerosis.Nature. 2011; 473: 317-325Crossref PubMed Scopus (2645) Google Scholar). Elevated LDL level is a leading risk factor for atherosclerosis (3Tsimikas S. Witztum J.L. The role of oxidized phospholipids in mediating lipoprotein(a) atherogenicity.Curr. Opin. Lipidol. 2008; 19: 369-377Crossref PubMed Scopus (99) Google Scholar). The progressive accumulation of LDL in the vessel wall drives the development of atherosclerosis (4Williams K.J. Tabas I. The response-to-retention hypothesis of early atherogenesis.Arterioscler. Thromb. Vasc. Biol. 1995; 15: 551-561Crossref PubMed Google Scholar). The initiation of atherosclerosis may be mediated by the subendothelial retention of LDL (5Nakashima Y. Fujii H. Sumiyoshi S. Wight T.N. Sueishi K. Early human atherosclerosis: accumulation of lipid and proteoglycans in intimal thickenings followed by macrophage infiltration.Arterioscler. Thromb. Vasc. Biol. 2007; 27: 1159-1165Crossref PubMed Scopus (316) Google Scholar, 6Tabas I. Williams K.J. Boren J. Subendothelial lipoprotein retention as the initiating process in atherosclerosis: update and therapeutic implications.Circulation. 2007; 116: 1832-1844Crossref PubMed Scopus (977) Google Scholar). This results from unbalanced dynamics of LDL, i.e., increased transfer to the arterial wall and retention by the extracellular matrix, mainly the proteoglycans (7Proctor S.D. Vine D.F. Mamo J.C. Arterial retention of apolipoprotein B(48)- and B(100)-containing lipoproteins in atherogenesis.Curr. Opin. Lipidol. 2002; 13: 461-470Crossref PubMed Scopus (163) Google Scholar, 8Nordestgaard B.G. Nielsen L.B. Atherosclerosis and arterial influx of lipoproteins.Curr. Opin. Lipidol. 1994; 5: 252-257Crossref PubMed Scopus (101) Google Scholar). For instance, in rabbit atherosclerosis models, injected LDL accumulated focally at atherosclerosis-prone regions of the arterial wall (9Nievelstein P.F. Fogelman A.M. Mottino G. Frank J.S. Lipid accumulation in rabbit aortic intima 2 hours after bolus infusion of low density lipoprotein. A deep-etch and immunolocalization study of ultrarapidly frozen tissue.Arterioscler. Thromb. 1991; 11: 1795-1805Crossref PubMed Google Scholar, 10Schwenke D.C. Carew T.E. Initiation of atherosclerotic lesions in cholesterol-fed rabbits. I. Focal increases in arterial LDL concentration precede development of fatty streak lesions.Arteriosclerosis. 1989; 9: 895-907Crossref PubMed Google Scholar). Similarly, transgenic mice expressing proteoglycan-binding defective LDLs exhibit a significantly lower rate of atherosclerosis compared with mice expressing WT LDL (11Skålén K. Gustafsson M. Rydberg E.K. Hulten L.M. Wiklund O. Innerarity T.L. Boren J. Subendothelial retention of atherogenic lipoproteins in early atherosclerosis.Nature. 2002; 417: 750-754Crossref PubMed Scopus (728) Google Scholar). Proteoglycans are major components of the extracellular matrix lining the arterial wall (12Camejo G. Hurt-Camejo E. Wiklund O. Bondjers G. Association of apo B lipoproteins with arterial proteoglycans: pathological significance and molecular basis.Atherosclerosis. 1998; 139: 205-222Abstract Full Text Full Text PDF PubMed Scopus (290) Google Scholar, 13Tannock L.R. King V.L. Proteoglycan mediated lipoprotein retention: a mechanism of diabetic atherosclerosis.Rev. Endocr. Metab. Disord. 2008; 9: 289-300Crossref PubMed Scopus (40) Google Scholar). Typically, proteoglycans consist of a core protein and one or multiple covalently linked glycosaminoglycans (GAGs) (14Varki A. Cummings R. Esko J. Freeze H. Hart G. Marth J. Essentials of Glycobiology.Cold Spring Harbor. Cold Spring Harbor Lab Press, New York1999Google Scholar). The proteoglycan, perlecan, is normally synthesized by endothelial cells, before being deposited in the subendothelial extracellular matrix (15Iozzo R.V. Basement membrane proteoglycans: from cellar to ceiling.Nat. Rev. Mol. Cell Biol. 2005; 6: 646-656Crossref PubMed Scopus (398) Google Scholar). Perlecan consists of a core protein with a molecular mass of ∼450 kDa and three long side chains of heparan sulfate (HS), which are critical in atherosclerosis development (16Tran-Lundmark K. Tran P.K. Paulsson-Berne G. Friden V. Soininen R. Tryggvason K. Wight T.N. Kinsella M.G. Boren J. Hedin U. Heparan sulfate in perlecan promotes mouse atherosclerosis: roles in lipid permeability, lipid retention, and smooth muscle cell proliferation.Circ. Res. 2008; 103: 43-52Crossref PubMed Google Scholar). The core protein has five domains (DI–DV) with disparate The contains sites for HS side chains. is to that DII contains repeat units that are highly homologous to the ligand-binding of LDL receptor (LDLR). Perlecan is present in the lesions of or mice of and perlecan, but in lesions of of Thromb. Vasc. Biol. 2002; PubMed Scopus Google and its with In advanced lesions with lipid the level of perlecan increased Y. G. Y. E. Y. Atherosclerosis in perlecan Lipid Res. Full Text Full Text PDF PubMed Scopus Google Scholar). with a of perlecan a of perlecan in the arterial wall and the in atherosclerosis in mice Y. G. Y. E. Y. Atherosclerosis in perlecan Lipid Res. Full Text Full Text PDF PubMed Scopus Google Scholar). Perlecan may to atherosclerosis its interaction with (12Camejo G. Hurt-Camejo E. Wiklund O. Bondjers G. Association of apo B lipoproteins with arterial proteoglycans: pathological significance and molecular basis.Atherosclerosis. 1998; 139: 205-222Abstract Full Text Full Text PDF PubMed Scopus (290) Google Scholar, K. Tran P.K. Paulsson-Berne G. Friden V. Soininen R. Tryggvason K. Wight T.N. Kinsella M.G. Boren J. Hedin U. Heparan sulfate in perlecan promotes mouse atherosclerosis: roles in lipid permeability, lipid retention, and smooth muscle cell proliferation.Circ. Res. 2008; 103: 43-52Crossref PubMed Google Scholar). The perlecan HS LDL and promotes LDL of perlecan HS in mice significantly atherosclerosis (16Tran-Lundmark K. Tran P.K. Paulsson-Berne G. Friden V. Soininen R. Tryggvason K. Wight T.N. Kinsella M.G. Boren J. Hedin U. Heparan sulfate in perlecan promotes mouse atherosclerosis: roles in lipid permeability, lipid retention, and smooth muscle cell proliferation.Circ. Res. 2008; 103: 43-52Crossref PubMed Google Scholar). of mouse perlecan that DII a domain with a and is M. K. Y. R. of domain of the membrane proteoglycan PubMed Scopus Google Scholar). the of perlecan DII in LDL has been In this we demonstrate that the core protein of perlecan interacts with LDL its LDLR-like DII. We that the secreted DII with O-linked glycans and its interaction with LDL largely on the glycosylation. Interestingly, the glycans sialic that are critical for the We that perlecan and its sialic are in human atherosclerotic arterial wall. study that perlecan sialic glycosylation to the development of atherosclerosis at the early and smooth muscle in with and and in with and For secreted protein in in with cell by in a cell and in the of for perlecan DII, and by the The DII, and by for by with or multiple to to and the and of DII. The at For the domain of human the by with an as a a from A. and of a of the of for with sialic Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). The at A the from synthesized and at sites M. K. Y. R. of domain of the membrane proteoglycan PubMed Scopus Google Scholar). The as or to the with or with as in the and for with or or For the in and at for with in the but with and for The in this study and from three with and for after with perlecan DII, and from cells, or The perlecan DII from For secreted from by Cell by in the The and cell with or in and at The by and three with and three with the The with the a and by protein with for or in at The by and with and the The with human LDL or in a and or in with at for The by and three with the The with the and with on a at the as the with the the to the as before M. S. E.J. R. of in and for of protein and PubMed Scopus Google Scholar). in injected the for which in and A in the perlecan DII injected followed by at at from to The and to a The O-linked glycans of WT perlecan DII from from Biol. Full Text PDF PubMed Google Scholar). The by through by to in as P. Y. I. of glycans for mass 2005; 19: PubMed Scopus Google Scholar). of by with and infusion a mass For cell with or perlecan DII in the with as S. A. S. M. of receptor as a density lipoprotein PubMed Scopus Google Scholar). the with and with as of glycosylation and protein Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). For human atherosclerosis and frozen with the or mouse and the in the with and in the in the with a with The and in the at the Perlecan DII has five units with of to the ligand-binding of (15Iozzo R.V. Basement membrane proteoglycans: from cellar to ceiling.Nat. Rev. Mol. Cell Biol. 2005; 6: 646-656Crossref PubMed Scopus (398) Google In of this the of perlecan with LDL has been this we a of the perlecan DII with an for and protein in and cells, and the protein present in the DII a significantly as compared with the DII a The of and is in the of DII with human LDL, we in and The of with LDL significantly greater than 2 and The with the and as a A but with an only for but for and The results that the perlecan DII interacts with LDL and the interaction is largely on the the interaction at the cell we of cells, which are S. A. S. M. of receptor as a density lipoprotein PubMed Scopus Google Scholar). We or DII the The with and with of perlecan DII but the the retention of at the cell that the interaction at the cell the the we DII and it in with the results the of than that of 2 with The and in for its and with We a rate and a rate The the DII and Interestingly, the at the condition is to that of with LDL as P. S. M. J.C. D. Cummings A. et of and low density lipoprotein on LDL receptor Biol. 2007; Full Text Full Text PDF PubMed Scopus Google Scholar). the of perlecan DII with LDL, we an in The results that the LDL to LDL the results that the perlecan DII in LDL which is largely on its We to the and functional of perlecan DII of the with to and glycans in a with which of the protein any glycosylation be any other but it multiple sites that the be O-linked glycosylation. The DII in WT and cell The of from which are defective in glycosylation H. J. in is to a in Biol. Full Text Full Text PDF PubMed Scopus Google to that from WT that DII The of DII in and which are defective in sialic M. R. of the the Biol. 1998; Full Text Full Text PDF PubMed Scopus Google and and which are defective in sialic and S. P. R. in glycosylation that the and Biol. Full Text Full Text PDF PubMed Scopus Google that it contains sialic and further perlecan DII glycosylation repeat as the DII The results in show a for the secreted repeat 2 with and a for the secreted repeat with as compared with that are and the glycans from repeat 2 to the of the secreted DII. In and have the and secreted with with and with that are the second of DII is significantly from a and contains no but is rich in In with this mouse perlecan has a unit, which is M. K. Y. R. of domain of the membrane proteoglycan PubMed Scopus Google Scholar). the glycosylation sites in 2 and we a of we that the with of and from repeat 2 that sites are on this we more for any we that of and from repeat and or serine from repeat with of in with and with and the of a secreted is to the WT with that the is defective in glycosylation. The glycosylation sites are in which are to the glycosylation in DII of mouse perlecan M. K. Y. R. of domain of the membrane proteoglycan PubMed Scopus Google Scholar). further the we of the DII We and the secreted from with the WT DII and for the in with human The results that the the of with which is to the and 2 with and that the is in binding. The HS side chains on perlecan with LDL (16Tran-Lundmark K. Tran P.K. Paulsson-Berne G. Friden V. Soininen R. Tryggvason K. Wight T.N. Kinsella M.G. Boren J. Hedin U. Heparan sulfate in perlecan promotes mouse atherosclerosis: roles in lipid permeability, lipid retention, and smooth muscle cell proliferation.Circ. Res. 2008; 103: 43-52Crossref PubMed Google and in this study we demonstrate that DII interacts with LDL. any DII and in LDL we that only and The and as as WT DII in WT and in G. A. Esko J.D. cell defective in and Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). The of the that the and from WT with but from the and and the multiple of protein the of HS side chains The DII from WT and have no and of the for the in with human protein the The DII from WT and show which is than WT but and with the of the from WT is than WT DII or and with and for The of WT with largely on the of the of the from the the with Similarly, the HS of the from the its to the level of WT DII with The results the that be an additive effect perlecan DII and on binding. results from the in We a to further the additive We cell expressing WT DII, and from WT The cell for with LDL. The results show that of and WT DII increased the by and of increased The results are with the from the in that be a perlecan HS on DII and in with The results the role of the O-linked glycans in mediating the interaction of perlecan DII with LDL. We the of the glycans of the WT and DII with the or a of WT protein by the for for for for and sialic or for The results demonstrate that the glycans of DII and Importantly, WT DII by which is for sialic in an but by a for sialic in an of the the that it the we mass to the of the The most five with multiple are in feature from structures is that most of the glycans the sialic This is with the of DII in and and the with the sialic that sialic significantly to the O-linked through J.C. of with a 2008; PubMed Scopus Google Scholar, H. of mass and 2014; PubMed Scopus Google Scholar). that the a role in mediating the interaction of DII with LDL, we of to the from the with significantly the of the WT protein with the sialic be by the from the WT that the of the sialic with The or WT and and the for in with human LDL. The LDL with with the of sialic the with LDL by with and for results that the perlecan DII contains sialic which the interaction of DII with LDL. The from the and to an for the sialic in the development of atherosclerosis. and atherosclerotic arterial by with a and the Perlecan and the sialic at low the arterial wall with the most in the membrane In in the atherosclerotic arterial perlecan highly in of lipid accumulation in the arterial The from sialic and with perlecan The sialic in and atherosclerotic further the sialic be with the accumulation in the we the with the and the We in the at the to the side of which may the of LDL from The accumulation with the sialic at the rich in In the atherosclerotic the more the arterial and its with the that the sialic with that perlecan and its sialic be in the development of atherosclerosis. Previous studies have linked perlecan with atherosclerosis. Perlecan is in the atherosclerotic lesions and its with that the role of perlecan in atherosclerosis is on the interaction the negatively charged HS on and the charged domains of (16Tran-Lundmark K. Tran P.K. Paulsson-Berne G. Friden V. Soininen R. Tryggvason K. Wight T.N. Kinsella M.G. Boren J. Hedin U. Heparan sulfate in perlecan promotes mouse atherosclerosis: roles in lipid permeability, lipid retention, and smooth muscle cell proliferation.Circ. Res. 2008; 103: 43-52Crossref PubMed Google Scholar). Perlecan contains LDLR-like DII. et M. K. Y. R. of domain of the membrane proteoglycan PubMed Scopus Google that the domain contains multiple O-linked glycosylation the of perlecan DII and its glycosylation in LDL has been In this we demonstrate that the core protein interacts with LDL its LDLR-like DII and the interaction is largely on the O-linked glycans only present in the secreted DII. We that most of the glycans are capped with the negatively charged sialic in which are for the in LDL binding. We further that the of perlecan and sialic is in the human atherosclerotic that the O-linked glycans be with the development of atherosclerosis. The that the perlecan DII interacts with LDL that the core protein may play a role in the LDL retention in the arterial wall that The it for the arterial perlecan to with the LDLs that through the endothelial from the and the interaction may the subendothelial LDL has been that the subendothelial retention of LDL is a critical in the development of atherosclerosis I. Williams K.J. Boren J. Subendothelial lipoprotein retention as the initiating process in atherosclerosis: update and therapeutic implications.Circulation. 2007; 116: 1832-1844Crossref PubMed Scopus (977) Google Scholar). For and of it that the of atherosclerosis in a of fatty streak that from the of lipoproteins (5Nakashima Y. Fujii H. Sumiyoshi S. Wight T.N. Sueishi K. Early human atherosclerosis: accumulation of lipid and proteoglycans in intimal thickenings followed by macrophage infiltration.Arterioscler. Thromb. Vasc. Biol. 2007; 27: 1159-1165Crossref PubMed Scopus (316) Google Scholar). The than the of in the early studies have that the perlecan and as as with LDL (16Tran-Lundmark K. Tran P.K. Paulsson-Berne G. Friden V. Soininen R. Tryggvason K. Wight T.N. Kinsella M.G. Boren J. Hedin U. Heparan sulfate in perlecan promotes mouse atherosclerosis: roles in lipid permeability, lipid retention, and smooth muscle cell proliferation.Circ. Res. 2008; 103: 43-52Crossref PubMed Google Scholar, Gustafsson M. Boren J. of the proteoglycan in apolipoprotein Biol. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, V. M. Hurt-Camejo E. J. smooth muscle LDL through LDL protein and LDL Thromb. Vasc. Biol. 2002; PubMed Scopus Google Scholar). to the proteoglycans is that The interaction the negatively charged and the charged domains of to be the molecular for the lipoprotein The of the perlecan LDLR-like DII from this study a LDL as compared with the interaction the and LDL. The perlecan domain may a of LDLR-like perlecan contains only the LDLR-like ligand-binding the that are for the LDL and as a G. K. K. J.L. J. of the LDL receptor extracellular domain at 2002; PubMed Scopus Google Scholar). Thus, the of the domain with LDL and its effect with HS on LDL the LDL subendothelial retention in the arterial wall. Perlecan DII that LDL binding. Among its five are homologous to the LDL of LDLR, and repeat units the for For of the has to three and a of negatively charged any to the which are to the charged of The contains such and the and functional demonstrate that are for the LDL and of any of the five the J.L. of of low density lipoprotein receptor to Biol. 1989; Full Text PDF PubMed Google Scholar). The second of perlecan DII is highly are no cysteine and negatively charged this repeat the of the The of such a repeat DII to be functional in LDL binding. the DII and the have a in LDL binding. We that the secreted domain interacts with LDL in a Interestingly, most of the glycosylation sites of DII from the second that is rich in A for the glycosylation is that most of the O-linked glycans are capped with sialic acids, which are to be negatively charged A. 2007; PubMed Scopus Google Scholar). The further by the of the sialic acids, which the binding. This is with the critical role of the of negatively charged of in LDL binding. of the the LDL The role of the is in the of the repeat units H. and of the lipoprotein Rev. 2005; PubMed Scopus Google Scholar). The are on the of the repeat that with the on the of the results from this study that the sialic glycans be to the of the for this to be that the sialic but sialic is in the The may be linked to the or sialic of and human and sialic acids, that are in in the but with a R. of sialic and A of and in J. 2011; PubMed Scopus Google Scholar, of human and sialic in the Res. 6: PubMed Scopus Google Scholar). In this perlecan DII is with the sialic in it to be is any sialic for the The interaction The most one is that the interaction which is from the perlecan is an extracellular matrix to the extracellular is of the of acids, with a portion of S. of the of 1991; Full Text PDF PubMed Scopus Google Scholar). In as for perlecan DII, the may be to with this it that the interaction of human aortic proteoglycans with LDL is by M. K. in of and oxidized low density lipoprotein to human aortic Biol. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar). of the of perlecan and its sialic is with the that of proteoglycans may the of atherosclerosis I. Williams K.J. Boren J. Subendothelial lipoprotein retention as the initiating process in atherosclerosis: update and therapeutic implications.Circulation. 2007; 116: 1832-1844Crossref PubMed Scopus (977) Google Scholar). Perlecan to be in the early in and mouse and its with the atherogenic of and perlecan, but in lesions of of Thromb. Vasc. Biol. 2002; PubMed Scopus Google Scholar, Y. G. Y. E. Y. Atherosclerosis in perlecan Lipid Res. Full Text Full Text PDF PubMed Scopus Google Scholar). study that it is in human atherosclerotic The perlecan is synthesized from the or smooth muscle at the G. G. Hurt-Camejo E. Bondjers G. of low density lipoproteins by proteoglycans synthesized by and human arterial smooth muscle Biol. Full Text PDF PubMed Google Scholar). is that oxidized LDL is to endothelial to that and smooth muscle to the arterial wall S.D. M. R. Fogelman A.M. low density lipoprotein induces protein in human endothelial and smooth muscle PubMed Scopus Google Scholar). The oxidized LDL the oxidized LDL, is to S. D. low density a role in and retention of during PubMed Scopus Google Scholar, I. O. lipoprotein is for arterial smooth muscle in PubMed Scopus Google Scholar). it that the secreted the proteoglycans sulfate but only a of HS leading to lipoprotein retention and the initiation of Thromb. Vasc. Biol. PubMed Scopus Google Scholar). we that the perlecan is most from the smooth muscle that the proteoglycans synthesized from the lesions more atherogenic This study that perlecan is with the of sialic O-linked which in the interaction with LDL. In that the perlecan core protein interacts with LDL. We show that the interaction is mediated by its LDLR-like DII in a on sialic Thus, the perlecan sialic is with the development of atherosclerosis and further study is to this The E. and for to this for the and and for in the The to and for the and for cells, and for the with domain domains heparan sulfate LDL receptor secreted domain

Evidence for a Structural Role for Acid-Fast Lipids in Oocyst Walls of <i>Cryptosporidium</i> , <i>Toxoplasma</i> , and <i>Eimeria</i>

mBio · 2013-09-04 · 83 citations

UNLABELLED: Coccidia are protozoan parasites that cause significant human disease and are of major agricultural importance. Cryptosporidium spp. cause diarrhea in humans and animals, while Toxoplasma causes disseminated infections in fetuses and untreated AIDS patients. Eimeria is a major pathogen of commercial chickens. Oocysts, which are the infectious form of Cryptosporidium and Eimeria and one of two infectious forms of Toxoplasma (the other is tissue cysts in undercooked meat), have a multilayered wall. Recently we showed that the inner layer of the oocyst walls of Toxoplasma and Eimeria is a porous scaffold of fibers of β-1,3-glucan, which are also present in fungal walls but are absent from Cryptosporidium oocyst walls. Here we present evidence for a structural role for lipids in the oocyst walls of Cryptosporidium, Toxoplasma, and Eimeria. Briefly, oocyst walls of each organism label with acid-fast stains that bind to lipids in the walls of mycobacteria. Polyketide synthases similar to those that make mycobacterial wall lipids are abundant in oocysts of Toxoplasma and Eimeria and are predicted in Cryptosporidium. The outer layer of oocyst wall of Eimeria and the entire oocyst wall of Cryptosporidium are dissolved by organic solvents. Oocyst wall lipids are complex mixtures of triglycerides, some of which contain polyhydroxy fatty acyl chains like those present in plant cutin or elongated fatty acyl chains like mycolic acids. We propose a two-layered model of the oocyst wall (glucan and acid-fast lipids) that resembles the two-layered walls of mycobacteria (peptidoglycan and acid-fast lipids) and plants (cellulose and cutin). IMPORTANCE: Oocysts, which are essential for the fecal-oral spread of coccidia, have a wall that is thought responsible for their survival in the environment and for their transit through the stomach and small intestine. While oocyst walls of Toxoplasma and Eimeria are strengthened by a porous scaffold of fibrils of β-1,3-glucan and by proteins cross-linked by dityrosines, both are absent from walls of Cryptosporidium. We show here that all oocyst walls are acid fast, have a rigid bilayer, dissolve in organic solvents, and contain a complex set of triglycerides rich in polyhydroxy and long fatty acyl chains that might be synthesized by an abundant polyketide synthase. These results suggest the possibility that coccidia build a waxy coat of acid-fast lipids in the oocyst wall that makes them resistant to environmental stress.

Strategies To Discover the Structural Components of Cyst and Oocyst Walls

Eukaryotic Cell · 2013-10-05 · 85 citations

Cysts of Giardia lamblia and Entamoeba histolytica and oocysts of Toxoplasma gondii and Cryptosporidium parvum are the infectious and sometimes diagnostic forms of these parasites. To discover the structural components of cyst and oocyst walls, we have developed strategies based upon a few simple assumptions. Briefly, the most abundant wall proteins are identified by monoclonal antibodies or mass spectrometry. Structural components include a sugar polysaccharide (chitin for Entamoeba, β-1,3-linked glucose for Toxoplasma, and β-1,3-linked GalNAc for Giardia) and/or acid-fast lipids (Toxoplasma and Cryptosporidium). Because Entamoeba cysts and Toxoplasma oocysts are difficult to obtain, studies of walls of nonhuman pathogens (E. invadens and Eimeria, respectively) accelerate discovery. Biochemical methods to dissect fungal walls work well for cyst and oocyst walls, although the results are often unexpected. For example, echinocandins, which inhibit glucan synthases and kill fungi, arrest the development of oocyst walls and block their release into the intestinal lumen. Candida walls are coated with mannans, while Entamoeba cysts are coated in a dextran-like glucose polymer. Models for cyst and oocyst walls derive from their structural components and organization within the wall. Cyst walls are composed of chitin fibrils and lectins that bind chitin (Entamoeba) or fibrils of the β-1,3-GalNAc polymer and lectins that bind the polymer (Giardia). Oocyst walls of Toxoplasma have two distinct layers that resemble those of fungi (β-1,3-glucan in the inner layer) or mycobacteria (acid-fast lipids in the outer layer). Oocyst walls of Cryptosporidium have a rigid bilayer of acid-fast lipids and inner layer of oocyst wall proteins.

Glycoproteomic characterization of Trichomonas vaginalis

2012-01-01

β-1,3-Glucan, Which Can Be Targeted by Drugs, Forms a Trabecular Scaffold in the Oocyst Walls of<i>Toxoplasma</i>and<i>Eimeria</i>

mBio · 2012-09-26 · 41 citations

UNLABELLED: The walls of infectious pathogens, which are essential for transmission, pathogenesis, and diagnosis, contain sugar polymers that are defining structural features, e.g., β-1,3-glucan and chitin in fungi, chitin in Entamoeba cysts, β-1,3-GalNAc in Giardia cysts, and peptidoglycans in bacteria. The goal here was to determine in which of three walled forms of Toxoplasma gondii (oocyst, sporocyst, or tissue cyst) is β-1,3-glucan, the product of glucan synthases and glucan hydrolases predicted by whole-genome sequences of the parasite. The three most important discoveries were as follows. (i) β-1,3-glucan is present in oocyst walls of Toxoplasma and Eimeria (a chicken parasite that is a model for intestinal stages of Toxoplasma) but is absent from sporocyst and tissue cyst walls. (ii) Fibrils of β-1,3-glucan are part of a trabecular scaffold in the inner layer of the oocyst wall, which also includes a glucan hydrolase that has a novel glucan-binding domain. (iii) Echinocandins, which target the glucan synthase and kill fungi, arrest development of the Eimeria oocyst wall and prevent release of the parasites into the intestinal lumen. In summary, β-1,3-glucan, which can be targeted by drugs, is an important component of oocyst walls of Toxoplasma but is not a component of sporocyst and tissue cyst walls. IMPORTANCE: We show here that walls of Toxoplasma oocysts, the infectious stage shed by cats, contain β-1,3-glucan, a sugar polymer that is a major component of fungal walls. In contrast to fungi, β-1,3-glucan is part of a trabecular scaffold in the inner layer of the oocyst wall that is independent of the permeability barrier formed by the outer layer of the wall. While glucan synthase inhibitors kill fungi, these inhibitors arrest the development of the oocyst walls of Eimeria (an important chicken pathogen that is a surrogate for Toxoplasma) and block release of oocysts into the intestinal lumen. The absence of β-1,3-glucan in tissue cysts of Toxoplasma suggests that drugs targeted at the glucan synthase might be used to treat Eimeria in chickens but not to treat Toxoplasma in people.

The Glycoproteins of Giardia

2011-01-01

The Jacob2 Lectin of the Entamoeba histolytica Cyst Wall Binds Chitin and Is Polymorphic

PLoS neglected tropical diseases · 2010-07-20 · 24 citations

BACKGROUND: The infectious and diagnostic form of Entamoeba histolytica (Eh), cause of amebic dysentery and liver abscess, is the quadranucleate cyst. The cyst wall of Entamoeba invadens (Ei), a model for Eh, is composed of chitin fibrils and three sets of chitin-binding lectins that cross-link chitin fibrils (multivalent Jacob lectins), self-aggregate (Jessie lectins), and remodel chitin (chitinase). The goal here was to determine how well the Ei model applies to Entamoeba cysts from humans. METHODS/RESULTS: An Eh Jacob lectin (EhJacob2) has three predicted chitin-binding domains surrounding a large, Ser-rich spacer. Recombinant EhJacob2 made in transfected Eh trophozoites binds to particulate chitin. Sequences of PCR products using primers flanking the highly polymorphic spacer of EhJacob2 may be used to distinguish Entamoeba isolates. Antibodies to the EhJacob2, EhJessie3, and chitinase each recognize cyst walls of clinical isolates of Entamoeba. While numerous sera from patients with amebic intestinal infections and liver abscess recognize recombinant EhJacob1 and EhJessie3 lectins, few of these sera recognize recombinant EhJacob2. CONCLUSIONS/SIGNIFICANCE: The EhJacob2 lectin binds chitin and is polymorphic, and Jacob2, Jessie3, and chitinase are present in cyst walls of clinical isolates of Entamoeba. These results suggest there are substantial similarities between cysts of the human pathogen (Eh) and the in vitro model (Ei), even though there are quantitative and qualitative differences in their chitin-binding lectins.