Received May 09, 2009; Revised July 30, 2009; Accepted July 30, 2009.
Abstract
Trypanosoma brucei, a protozoan parasite, causes sleeping sickness in humans and Nagana disease in domestic animals in central Africa. The trypanosome surface is extensively covered by glycosylphosphatidylinositol (GPI)-anchored proteins known as variant surface glycoproteins and procyclins. GPI anchoring is suggested to be important for trypanosome survival and establishment of infection. Trypanosomes are not only pathogenically important, but also constitute a useful model for elucidating the GPI biosynthesis pathway. This review focuses on the trypanosome GPI biosynthesis pathway. Studies on GPI that will be described indicate the potential for the design of drugs that specifically inhibit trypanosome GPI biosynthesis.
Acosta-Serrano A,Vassella E,Liniger M,Kunz Renggli C,Brun R,Roditi I,Englund PT. The surface coat of procyclic Trypanosoma brucei: programmed expression and proteolytic cleavage of procyclin in the tsetse fly. Proc Natl Acad Sci USA 2001;98:1513–1518.
3.
Ferguson MA,Homans SW,Dwek RA,Rademacher TW. Glycosyl-phosphatidylinositol moiety that anchors Trypanosoma brucei variant surface glycoprotein to the membrane. Science 1988;239:753–759.
4.
Homans SW,Ferguson MA,Dwek RA,Rademacher TW,Anand R,Williams AF. Complete structure of the glycosyl phosphatidylinositol membrane anchor of rat brain Thy-1 glycoprotein. Nature 1988;333:269–272.
5.
Field MC,Menon AK,Cross GA. A glycosylphosphatidylinositol protein anchor from procyclic stage Trypanosoma brucei: lipid structure and biosynthesis. EMBO J 1991;10:2731–2739.
6.
Ferguson MAJ. Site of palmitoylation of a phospholipase C-resistant glycosyl-phosphatidylinositol membrane anchor. Biochem J 1992;284:297–300.
7.
Ferguson MAJ,Murray P,Rutherford H,McConville MJ. A simple purification of procyclic acidic repetitive protein and demonstration of a sialylated glycosyl-phosphatidylinositol membrane anchor. Biochem J 1993;291:51–55.
8.
Cross GA. Identification, purification and properties of clone-specific glycoprotein antigens constituting the surface coat of Trypanosoma brucei. Parasitology 1975;71:393–417.
9.
McConville MJ,Ferguson MAJ. The structure, biosynthesis and function of glycosylated phosphatidylinositols in the parasitic protozoa and higher eukaryotes. Biochem J 1993;294:305–324.
10.
Pays E,Nolan DP. Expression and function of surface proteins in Trypanosoma brucei. Mol Biochem Parasitol 1998;91:3–36.
11.
Clayton CE,Mowatt MR. The procyclic acidic repetitive proteins of Trypanosoma brucei. Purification and post-translational modification. J Biol Chem 1989;264:15088–15093.
12.
Ferguson MA,Murray P,Rutherford H,McConville MJ. A simple purification of procyclic acidic repetitive protein and demonstration of a sialylated glycosyl-phosphatidylinositol membrane anchor. Biochem J 1993;291(Pt 1):51–55.
13.
Ferguson MA. The structure, biosynthesis and functions of glycosylphosphatidylinositol anchors, and the contributions of trypanosome research. J Cell Sci 1999;112:2799–2809.
14.
McConville MJ,Menon AK. Recent developments in the cell biology and biochemistry of glycosylphosphatidylinositol lipids. Mol Membr Biol 2000;17:1–16.
15.
Udenfriend S,Kodukula K. How glycosylphosphatidylinositol-anchored membrane proteins are made. Annu Rev Biochem 1995;64:563–591.
16.
Tiede A,Bastisch I,Schubert J,Orlean P,Schmidt RE. Biosynthesis of glycosylphosphatidylinositols in mammalian and unicellular microbes. Biol Chem 1999;380:503–523.
17.
Ikezawa H. Glycosylphosphatidylinositol (GPI)-anchored proteins. Biol Pharm Bull 2002;25:409–417.
18.
Watanabe R,Inoue N,Westfall B,Taron CH,Orlean P,Takeda J,Kinoshita T. The first step of glycosylphosphatidylinositol biosynthesis is mediated by a complex of PIG-A, PIG-H, PIG-C and GPI1. EMBO J 1998;17:877–885.
19.
Watanabe R,Murakami Y,Marmor MD,Inoue N,Maeda Y,Hino J,Kangawa K,Julius M,Kinoshita T. Initial enzyme for glycosylphosphatidylinositol biosynthesis requires PIG-P and is regulated by DPM2. EMBO J 2000;19:4402–4411.
20.
Murakami Y,Siripanyaphinyo U,Hong Y,Tashima Y,Maeda Y,Kinoshita T. The initial enzyme for glycosylphosphatidylinositol biosynthesis requires PIG-Y, a seventh component. Mol Biol Cell 2005;16:5236–5246.
21.
Kawagoe K,Takeda J,Endo Y,Kinoshita T. Molecular cloning of murine Pig-a, a gene for GPI-anchor biosynthesis, and demonstration of interspecies conservation of its structure, function and gene locus. Genomics 1994;23:566–574.
22.
Nakamura N,Inoue N,Watanabe R,Takahashi M,Takeda J,Stevens VL,Kinoshita T. Expression cloning of PIG-L, a candidate N-acetylglucosaminyl-phosphatidylinositol deacetylase. J Biol Chem 1997;272:15834–15840.
23.
Chang T,Milne KG,Guther ML,Smith TK,Ferguson MA. Cloning of Trypanosoma brucei and Leishmania major genes encoding the GlcNAc-phosphatidylinositol de-N-acetylase of glycosylphosphatidylinositol biosynthesis that is essential to the African sleeping sickness parasite. J Biol Chem 2002;277:50176–50182.
24.
Urbaniak MD,Crossman A,Chang T,Smith TK,van Aalten DM,Ferguson MA. The N-acetyl-D-glucosaminylphosphatidylinositol de-N-acetylase of glycosylphosphatidylinositol biosynthesis is a zinc metalloenzyme. J Biol Chem 2005;280:22831–22838.
25.
Sharma DK,Smith TK,Weller CT,Crossman A,Brimacombe JS,Ferguson MAJ. Differences between the trypanosomal and human GlcNAc-PI de-N-acetylases of glycosylphosphatidylinositol membrane anchor biosynthesis. Glycobiology 1999;9:415–422.
26.
Smith TK,Crossman A,Borissow CN,Paterson MJ,Dix A,Brimacombe JS,Ferguson MA. Specificity of GlcNAc-PI de-N-acetylase of GPI biosynthesis and synthesis of parasite-specific suicide substrate inhibitors. EMBO J 2001;20:3322–3332.
27.
Murakami Y,Siripanyapinyo U,Hong Y,Kang JY,Ishihara S,Nakakuma H,Maeda Y,Kinoshita T. PIG-W is critical for inositol acylation but not for flipping of glycosylphosphatidylinositol-anchor. Mol Biol Cell 2003;14:4285–4295.
28.
Smith TK,Milne FC,Sharma DK,Crossman A,Brimacombe JS,Ferguson MA. Early steps in glycosylphosphatidylinositol biosynthesis in Leishmania major. Biochem J 1997;326:393–400.
29.
Güther ML,Ferguson MA. The role of inositol acylation and inositol deacylation in GPI biosynthesis in Trypanosoma brucei. EMBO J 1995;14:3080–3093.
30.
Urbaniak MD,Yashunsky DV,Crossman A,Nikolaev AV,Ferguson MA. Probing enzymes late in the trypanosomal glycosylphosphatidylinositol biosynthetic pathway with synthetic glycosylphosphatidylinositol analogues. ACS Chem Biol 2008;3:625–634.
31.
Güther ML,Masterson WJ,Ferguson MA. The effects of phenylmethylsulfonyl fluoride on inositol-acylation and fatty acid remodeling in African trypanosomes. J Biol Chem 1994;269:18694–18701.
32.
Guther ML,Ferguson MA. The role of inositol acylation and inositol deacylation in GPI biosynthesis in Trypanosoma brucei. EMBO J 1995;14:3080–3093.
33.
Güther ML,Masterson WJ,Ferguson MA. The role of glycolipid C in the GPI biosynthetic pathway in Trypanosoma brucei bloodstream forms. Braz J Med Biol Res 1994;27:121–126.
34.
Kajiwara K,Watanabe R,Pichler H,Ihara K,Murakami S,Riezman H,Funato K. Yeast ARV1 is required for efficient delivery of an early GPI intermediate to the first mannosyltransferase during GPI assembly and controls lipid flow from the endoplasmic reticulum. Mol Biol Cell 2008;19:2069–2082.
35.
Maeda Y,Watanabe R,Harris CL,Hong Y,Ohishi K,Kinoshita K,Kinoshita T. PIG-M transfers the first mannose to glycosylphosphatidylinositol on the lumenal side of the ER. EMBO J 2001;20:250–261.
36.
Ashida H,Hong Y,Murakami Y,Shishioh N,Sugimoto N,Kim YU,Maeda Y,Kinoshita T. Mammalian PIG-X and yeast Pbn1p are the essential components of glycosylphosphatidylinositol-mannosyltransferase I. Mol Biol Cell 2005;16:1439–1448.
37.
Smith TK,Sharma DK,Crossman A,Brimacombe JS,Ferguson MA. Selective inhibitors of the glycosylphosphatidylinositol biosynthetic pathway of Trypanosoma brucei. EMBO J 1999;18:5922–5930.
38.
Kang JY,Hong Y,Ashida H,Shishioh N,Murakami Y,Morita YS,Maeda Y,Kinoshita T. PIG-V involved in transferring the second mannose in glycosylphosphatidylinositol. J Biol Chem 2005;280:9489–9497.
39.
Takahashi M,Inoue N,Ohishi K,Maeda Y,Nakamura N,Endo Y,Fujita T,Takeda J,Kinoshita T. PIG-B, a membrane protein of the endoplasmic reticulum with a large lumenal domain, is involved in transferring the third mannose of the GPI anchor. EMBO J 1996;15:4254–4261.
40.
Nagamune K,Nozaki T,Maeda Y,Ohishi K,Fukuma T,Hara T,Schwarz RT,Sutterlin C,Brun R,Riezman H,Kinoshita T. Critical roles of glycosylphosphatidylinositol for Trypanosoma brucei. Proc Natl Acad Sci USA 2000;97:10336–10341.
41.
Hong Y,Maeda Y,Watanabe R,Inoue N,Ohishi K,Kinoshita T. Requirement of PIG-F and PIG-O for transferring phosphoethanolamine to the third mannose in glycosylphosphatidylinositol. J Biol Chem 2000;275:20911–20919.
42.
Hong Y,Maeda Y,Watanabe R,Ohishi K,Mishkind M,Riezman H,Kinoshita T. Pig-n, a mammalian homologue of yeast Mcd4p, is involved in transferring phosphoethanolamine to the first mannose of the glycosylphosphatidylinositol. J Biol Chem 1999;274:35099–35106.
43.
Shishioh N,Hong Y,Ohishi K,Ashida H,Maeda Y,Kinoshita T. GPI7 is the second partner of PIG-F and involved in modification of glycosylphosphatidylinositol. J Biol Chem 2005;280:9728–9734.
44.
Menon AK,Schwarz RT,Mayor S,Cross GA. Cell-free synthesis of glycosyl-phosphatidylinositol precursors for the glycolipid membrane anchor of Trypanosoma brucei variant surface glycoproteins. Structural characterization of putative biosynthetic intermediates. J Biol Chem 1990;265:9033–9042.
45.
Tanaka S,Maeda Y,Tashima Y,Kinoshita T. Inositol deacylation of glycosylphosphatidylinositol-anchored proteins is mediated by mammalian PGAP1 and yeast Bst1p. J Biol Chem 2004;279:14256–14263.
46.
Guther ML,Leal S,Morrice NA,Cross GA,Ferguson MA. Purification, cloning and characterization of a GPI inositol deacylase from Trypanosoma brucei. EMBO J 2001;20:4923–4934.
47.
Guther ML,Prescott AR,Ferguson MA. Deletion of the GPIdeAc gene alters the location and fate of glycosylphosphatidylinositol precursors in Trypanosoma brucei. Biochemistry 2003;42:14532–14540.
48.
Hong Y,Nagamune K,Morita YS,Nakatani F,Ashida H,Maeda Y,Kinoshita T. Removal or maintenance of inositol-linked acyl chain in GPI is critical in trypanosome life cycle. J Biol Chem 2006;281:11595–11602.
Guther ML,Masterson WJ,Ferguson MA. The effects of phenylmethylsulfonyl fluoride on inositol-acylation and fatty acid remodeling in African trypanosomes. J Biol Chem 1994;269:18694–18701.
51.
Guther ML,Masterson WJ,Ferguson MA. The role of glycolipid C in the GPI biosynthetic pathway in Trypanosoma brucei bloodstream forms. Braz J Med Biol Res 1994;27:121–126.
52.
Jaquenoud M,Pagac M,Signorell A,Benghezal M,Jelk J,Butikofer P,Conzelmann A. The Gup1 homologue of Trypanosoma brucei is a GPI glycosylphosphatidylinositol remodelase. Mol Microbiol 2008;67:202–212.
53.
Fujita M,Jigami Y. Lipid remodeling of GPI-anchored proteins and its function. Biochim Biophys Acta 2008;1780:410–420.
54.
Tashima Y,Taguchi R,Murata C,Ashida H,Kinoshita T,Maeda Y. PGAP2 is essential for correct processing and stable expression of GPI-anchored proteins. Mol Biol Cell 2006;17:1410–1420.
55.
Ohishi K,Inoue N,Kinoshita T. PIG-S and PIG-T, essential for GPI anchor attachment to proteins, form a complex with GAA1 and GPI8. EMBO J 2001;20:4088–4098.
56.
Hong Y,Ohishi K,Kang JY,Tanaka S,Inoue N,Nishimura J,Maeda Y,Kinoshita T. Human PIG-U and yeast Cdc91p are the fifth subunit of GPI transamidase that attaches GPI-anchors to proteins. Mol Biol Cell 2003;14:1780–1789.
57.
Nagamune K,Ohishi K,Ashida H,Hong Y,Hino J,Kangawa K,Inoue N,Maeda Y,Kinoshita T. GPI transamidase of Trypanosoma brucei has two previously uncharacterized (trypanosomatid transamidase 1 and 2) and three common subunits. Proc Natl Acad Sci USA 2003;100:10682–10687.
58.
Ohishi K,Nagamune K,Maeda Y,Kinoshita T. Two subunits of glycosylphosphatidylinositol transamidase, GPI8 and PIG-T, form a functionally important intermolecular disulfide bridge. J Biol Chem 2003;278:13959–13967.
59.
Ohishi K,Inoue N,Maeda Y,Takeda J,Riezman H,Kinoshita T. Gaa1p and gpi8p are components of a glycosylphosphatidylinositol (GPI) transamidase that mediates attachment of GPI to proteins. Mol Biol Cell 2000;11:1523–1533.
60.
Meyer U,Benghezal M,Imhof I,Conzelmann A. Active site determination of Gpi8p, a caspase-related enzyme required for glycosylphosphatidylinositol anchor addition to proteins. Biochemistry 2000;39:3461–3471.
61.
Vidugiriene J,Vainauskas S,Johnson AE,Menon AK. Endoplasmic reticulum proteins involved in glycosylphosphatidylinositol-anchor attachment: photocrosslinking studies in a cell-free system. Eur J Biochem 2001;268:2290–2300.
62.
Spurway TD,Dalley JA,High S,Bulleid NJ. Early events in glycosylphosphatidylinositol anchor addition: substrate proteins associates with the transamidase subunit Gpi8p. J Biol Chem 2001;276:15975–15982.
63.
Kang X,Szallies A,Rawer M,Echner H,Duszenko M. GPI anchor transamidase of Trypanosoma brucei: in vitro assay of the recombinant protein and VSG anchor exchange. J Cell Sci 2002;115:2529–2539.
64.
Lillico S,Field MC,Blundell P,Coombs GH,Mottram JC. Essential roles for GPI-anchored proteins in African trypanosomes revealed using mutants deficient in GPI8. Mol Biol Cell 2003;14:1182–1194.
65.
Benghezal M,Benachour A,Rusconi S,Aebi M,Conzelmann A. Yeast Gpi8p is essential for GPI anchor attachment onto proteins. EMBO J 1996;15:6575–6583.
66.
Hong Y,Nagamune K,Ohishi K,Morita YS,Ashida H,Maeda Y,Kinoshita T. TbGPI16 is an essential component of GPI transamidase in Trypanosoma brucei. FEBS Lett 2006;580:603–606.
67.
Low MG. The glycosyl-phosphatidylinositol anchor of membrane proteins. Biochim Biophys Acta 1989;988:427–454.
68.
Kinoshita T,Inoue N,Takeda J. Defective glycosyl phosphatidylinositol anchor synthesis and paroxysmal nocturnal hemoglobinuria. Adv Immunol 1995;60:57–103.
69.
Moran P,Caras IW. Requirements for glycosylphosphatidylinositol attachment are similar but not identical in mammalian cells and parasitic protozoa. J Cell Biol 1994;125:333–343.