Glycobiology of Giardia

Summary

Principal Investigator: John C Samuelson
Abstract: DESCRIPTION (provided by applicant): Giardia lamblia, which is spread by the fecal-oral route, is an important parasitic cause of diarrhea in the US and the developing world. The infectious and diagnostic stage of Giardia is the quadranucleate cyst, which has a wall composed in part by fibrils of a unique GalNAc homopolymer. During the previous funding period, we have made many discoveries concerning glycosylation of Giardia proteins (Aim 1) and the formation of the cyst wall (Aim 2). These discoveries have suggested two related hypotheses that will be pursued in the proposed experiments. First, protein glycosylation in Giardia is much simpler than that of the human host or of model eukaryotes such as Saccharomyces. Therefore studies of Giardia glycans in Aim 1 will provide insights concerning the diversity and evolution of protein glycosylation, as well as providing a better description of Giardia glycoproteins. The second hypothesis is that the cyst wall of Giardia is composed of a small number of proteins, which are lectins that bind fibrils of the GalNAc homopolymer. Studies in Aim 2 that characterize enzymes involved in synthesis and degradation of the Giardia cyst wall will lead to insights into the mechanism of pathogenesis of this parasite and provide an alternative wall model that is distinct from those of fungi that contain multiple sugar polymers and ~100 proteins. Progress towards Aim 1 included demonstration that Giardia makes a 2-sugar N-glycan precursor rather than the 14-sugar precursor made by metazoans and fungi. The Giardia oligosaccharyl transferase (OST) that transfers N-glycans has a single subunit rather than 8 subunits. There is selection for sites of N- linked glycans in eukaryotes with N-glycan-dependent quality control (QC) of protein folding, but there is no such selection in Giardia.UDP-GlcNAc is the only nucleotide-sugar transported into the ER lumen of Giardia. Proposed experiments towards Specific Aim 1 will characterize the single-subunit OST of Giardia and characterize what we predict is a unique GPI anchor of Giardia. Progress toward Aim 2 included use of wheat germ agglutinin, which binds to the Giardia N-glycans, to dramatically enrich secreted and membrane proteins. Cyst-specific glycoproteins included a candidate for the GalNAc homopolymer synthase. The GalNAc homopolymer, which was isolated free of protein, maintains its hollow spherical shape based upon interlocking loops of curled fibrils. The Leu-rich repeat domain of the Giardia cyst wall protein 1 is a lectin that binds the GalNAc homopolymer. Proposed experiments towards Specific Aim 1 will explore further a curled fibril and lectin model of the Giardia cyst wall, isolate the synthase for the unique GalNAc homopolymer, and characterize the glycohydrolase that breaks down the GalNAc polymer during excystation.
Funding Period: 2000-07-01 - 2016-04-30
more information: NIH RePORT

Top Publications

  1. pmc Sequence and structure evolved separately in a ribosomal ubiquitin variant
    Andre Catic
    Department of Biology, Whitehead Institute for Biomedical Research and Massachusetts Institute of Technology, Cambridge, MA 02142, USA
    EMBO J 26:3474-83. 2007
  2. pmc Evidence for a structural role for acid-fast lipids in oocyst walls of Cryptosporidium, Toxoplasma, and Eimeria
    G Guy Bushkin
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts, USA
    MBio 4:e00387-13. 2013
  3. pmc A simple fibril and lectin model for cyst walls of Entamoeba and perhaps Giardia
    John Samuelson
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA, USA
    Trends Parasitol 27:17-22. 2011
  4. pmc Giardia cyst wall protein 1 is a lectin that binds to curled fibrils of the GalNAc homopolymer
    Aparajita Chatterjee
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts, USA
    PLoS Pathog 6:e1001059. 2010
  5. pmc Trichomonas transmembrane cyclases result from massive gene duplication and concomitant development of pseudogenes
    Jike Cui
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts, USA
    PLoS Negl Trop Dis 4:e782. 2010
  6. pmc Molecular characterization of the cis-prenyltransferase of Giardia lamblia
    Kariona A Grabińska
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118, USA
    Glycobiology 20:824-32. 2010
  7. pmc Suggestive evidence for Darwinian Selection against asparagine-linked glycans of Plasmodium falciparum and Toxoplasma gondii
    G Guy Bushkin
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118, USA
    Eukaryot Cell 9:228-41. 2010
  8. pmc Darwinian selection for sites of Asn-linked glycosylation in phylogenetically disparate eukaryotes and viruses
    Jike Cui
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118, USA
    Proc Natl Acad Sci U S A 106:13421-6. 2009
  9. pmc Giardia, Entamoeba, and Trichomonas enzymes activate metronidazole (nitroreductases) and inactivate metronidazole (nitroimidazole reductases)
    Dibyarupa Pal
    Department of Biotechnology, Indian Institute of Technology, Kharagpur, West Bengal
    Antimicrob Agents Chemother 53:458-64. 2009
  10. pmc Molecular characterization of nucleocytosolic O-GlcNAc transferases of Giardia lamblia and Cryptosporidium parvum
    Sulagna Banerjee
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118, USA
    Glycobiology 19:331-6. 2009

Detail Information

Publications17

  1. pmc Sequence and structure evolved separately in a ribosomal ubiquitin variant
    Andre Catic
    Department of Biology, Whitehead Institute for Biomedical Research and Massachusetts Institute of Technology, Cambridge, MA 02142, USA
    EMBO J 26:3474-83. 2007
    ..Based on these observations, we propose a model to explain the origin of the widespread ubiquitin superfold in eukaryotes...
  2. pmc Evidence for a structural role for acid-fast lipids in oocyst walls of Cryptosporidium, Toxoplasma, and Eimeria
    G Guy Bushkin
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts, USA
    MBio 4:e00387-13. 2013
    ..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)...
  3. pmc A simple fibril and lectin model for cyst walls of Entamoeba and perhaps Giardia
    John Samuelson
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA, USA
    Trends Parasitol 27:17-22. 2011
    ..Although many of the details remain to be determined for the cyst wall of Giardia, current data suggest a relatively simple fibril and lectin model for the Entamoeba cyst wall...
  4. pmc Giardia cyst wall protein 1 is a lectin that binds to curled fibrils of the GalNAc homopolymer
    Aparajita Chatterjee
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts, USA
    PLoS Pathog 6:e1001059. 2010
    ..During excystation, host and Giardia proteases appear to degrade bound CWPs, exposing fibrils of the GalNAc homopolymer that are digested by a stage-specific glycohydrolase...
  5. pmc Trichomonas transmembrane cyclases result from massive gene duplication and concomitant development of pseudogenes
    Jike Cui
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts, USA
    PLoS Negl Trop Dis 4:e782. 2010
    ..With the goal of beginning to understand why some Trichomonas genes are present in so many copies, we characterized here a family of approximately 123 Trichomonas genes that encode transmembrane adenylyl cyclases (TMACs)...
  6. pmc Molecular characterization of the cis-prenyltransferase of Giardia lamblia
    Kariona A Grabińska
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118, USA
    Glycobiology 20:824-32. 2010
    ..These results suggest that the synthetic pathway for Dol-P is conserved in Giardia, even if some of the important enzymes are different from those of higher eukaryotes or remain unidentified...
  7. pmc Suggestive evidence for Darwinian Selection against asparagine-linked glycans of Plasmodium falciparum and Toxoplasma gondii
    G Guy Bushkin
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118, USA
    Eukaryot Cell 9:228-41. 2010
    ..In addition, occupation of N-glycan sites is markedly reduced in apicoplast proteins versus some secretory proteins in both Plasmodium and Toxoplasma...
  8. pmc Darwinian selection for sites of Asn-linked glycosylation in phylogenetically disparate eukaryotes and viruses
    Jike Cui
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118, USA
    Proc Natl Acad Sci U S A 106:13421-6. 2009
    ..We conclude that there is Darwinian selection for sequons in phylogenetically disparate eukaryotes and viruses...
  9. pmc Giardia, Entamoeba, and Trichomonas enzymes activate metronidazole (nitroreductases) and inactivate metronidazole (nitroimidazole reductases)
    Dibyarupa Pal
    Department of Biotechnology, Indian Institute of Technology, Kharagpur, West Bengal
    Antimicrob Agents Chemother 53:458-64. 2009
    ....
  10. pmc Molecular characterization of nucleocytosolic O-GlcNAc transferases of Giardia lamblia and Cryptosporidium parvum
    Sulagna Banerjee
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118, USA
    Glycobiology 19:331-6. 2009
    ..These results suggest the possibility that O-GlcNAc modification of nucleocytosolic proteins also has function(s) in simple eukaryotes...
  11. pmc Changes in the N-glycome, glycoproteins with Asn-linked glycans, of Giardia lamblia with differentiation from trophozoites to cysts
    Daniel M Ratner
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, 715 Albany Street, Evans 425, Boston, MA 02118, USA
    Eukaryot Cell 7:1930-40. 2008
    ..In summary, there are major changes in the Giardia N-glycome with the differentiation from trophozoites to cysts...
  12. pmc Dolichyl-phosphate-glucose is used to make O-glycans on glycoproteins of Trichomonas vaginalis
    Kariona A Grabińska
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, 715 Albany Street, Evans 425, Boston, MA 02118, USA
    Eukaryot Cell 7:1344-51. 2008
    ..To our knowledge, this is the first demonstration of Dol-P-Glc as a sugar donor for O-glycans on glycoproteins...
  13. pmc Use of Giardia, which appears to have a single nucleotide-sugar transporter for UDP-GlcNAc, to identify the UDP-Glc transporter of Entamoeba
    Sulagna Banerjee
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118 2492, USA
    Mol Biochem Parasitol 159:44-53. 2008
    ..In summary, all three NSTs of Entamoeba and the single NST of Giardia have been molecularly characterized, and transfected Giardia provides a new system for testing heterologous UDP-Glc transporters...
  14. ncbi A single Caenorhabditis elegans Golgi apparatus-type transporter of UDP-glucose, UDP-galactose, UDP-N-acetylglucosamine, and UDP-N-acetylgalactosamine
    Carolina E Caffaro
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts, USA
    Biochemistry 47:4337-44. 2008
    ..elegans have been found to transport two or three substrates, the substrate specificity of ZK896.9 raises questions as to the evolutionary ancestry of this group of proteins in this nematode...
  15. pmc The evolution of N-glycan-dependent endoplasmic reticulum quality control factors for glycoprotein folding and degradation
    Sulagna Banerjee
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, MA 02118, USA
    Proc Natl Acad Sci U S A 104:11676-81. 2007
    ..We conclude that N-glycan precursor length has profound effects on N-glycan-dependent QC of glycoprotein folding and ERAD...
  16. pmc Strategies to discover the structural components of cyst and oocyst walls
    John Samuelson
    Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts, USA
    Eukaryot Cell 12:1578-87. 2013
    ..Oocyst walls of Cryptosporidium have a rigid bilayer of acid-fast lipids and inner layer of oocyst wall proteins. ..

Research Grants30

  1. Phospholipid flip-flop in biogenic membranes
    Anant K Menon; Fiscal Year: 2013
    ..coli lipopolysaccharide. We are interested in identifying the transport proteins that catalyze lipid flipping in yeast and mammals and understanding how they work. ..
  2. Pacific Southwest RCE for Biodefense &Emerging Infectious Diseases Research
    Alan G Barbour; Fiscal Year: 2013
    ..abstract_text> ..
  3. Functional Glycomics of Human Saliva
    Susan J Fisher; Fiscal Year: 2013
    ..Additionally, competitive inhibitors could be used to disrupt the adhesion of disease-related organisms and, conversely, oligosaccharide sequences that support adhesion of beneficial species could be used to improve the oral ecology. ..
  4. Contribution of N-glycosylation to the Toxoplasma glycoproteome
    Anthony P Sinai; Fiscal Year: 2013
    ..This work represents the first in depth study of the cell biologic basis of protein glycosylation i Toxoplasma using state of the art approaches to modulate gene expression. ..
  5. Pacific NorthWest Regional Center of Excellence (PNWRCE)
    Jay A Nelson; Fiscal Year: 2013
    ..pseudomallei host pathogen response during both the septicemic as well as the intracellular phases of the disease. ..
  6. Southeast Regional Centers of Excellence for Biodefense &Emerging Infectious Di
    Philip Frederick Sparling; Fiscal Year: 2013
    ..SERCEB brings new investigators to the biodefense effort through a combination of educational programs, support of innovative new projects, and the synergistic interactions among its world-class investigators. ..
  7. Small molecule protein-glycan inhib. as malaria transmission-blocking therapuetic
    RHOEL DAVID RAMOS DINGLASAN; Fiscal Year: 2013
    ....
  8. LIAI Epitope Validation Center: Characterization of Allergen specific T Cells
    ALESSANDRO D SETTE; Fiscal Year: 2013
    ..abstract_text> ..
  9. Novel diagnostics and therapeutics for caliciviruses
    MARY KOLB ESTES; Fiscal Year: 2013
    ..abstract_text> ..
  10. Host glycosyltransferases in the glycosylation of Toxoplasma proteins
    Anthony P Sinai; Fiscal Year: 2013
    ....
  11. N-Linked Protein Glycosylation: Pathways and Processes
    Barbara Imperiali; Fiscal Year: 2013
    ....
  12. 2013/2015 Glycobiology Gordon Research Conference &Gordon Research Seminar
    Kelley W Moremen; Fiscal Year: 2013
    ..The meeting will focus on the latest hot-topic research areas at the frontiers of biomedical research and bring together the leading and developing scientists in the field. ..
  13. Photocrosslinking characterization of heat-labile toxin glycoreceptors
    Amberlyn M Wands; Fiscal Year: 2013
    ..These experiments will mark an important achievement in understanding the relevant proteins involved in ETEC mediated disease, and thus uncover novel therapeutic targets to complement existing treatments. ..