Giardia lamblia:Incorporation of Free and Conjugated Fatty Acids into Glycerol-Based Phospholipids

Gibson, George R.; Ramirez, David A.; Maier, Julie; Castillo, Cynthia; Das, Siddhartha

doi:10.1006/expr.1999.4389

Cited by 30 publications

(29 citation statements)

References 33 publications

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“…To investigate the nature of the fi rst N-terminal hydrophobic sequence, HA tags were introduced at the N-terminal end either of the full-length GlGCS or of a variant lacking the fi rst hydrophobic stretch (aa [3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22]. Both reporters showed an equivalent signal and distribution, indicating that, like the mammalian homolog, the parasite protein is not proteolytically processed to remove the fi rst hydrophobic sequence (data not shown).…”

Section: Discussionmentioning

confidence: 99%

“…Lipid neosynthesis is limited in G. lamblia , and the parasite is thought to rely on lipids taken up from the environment, namely, the host intestinal content. Indeed, in vitro analyses showed negligible incorporation of lipid precursors, such as acetate and glycerol ( 11 ), while robust incorporation of exogenous radiolabeled fatty acids (12)(13)(14), phospholipids ( 14 ), ceramide, and gangliosides ( 15,16 ) could be demonstrated. In addition, uptake of fl uorescent sphingolipid analogs, including ceramide and sphingomyelin, has been reported ( 12,14,15 ).…”

Section: Gene Expression Analysismentioning

confidence: 99%

“…Indeed, in vitro analyses showed negligible incorporation of lipid precursors, such as acetate and glycerol ( 11 ), while robust incorporation of exogenous radiolabeled fatty acids (12)(13)(14), phospholipids ( 14 ), ceramide, and gangliosides ( 15,16 ) could be demonstrated. In addition, uptake of fl uorescent sphingolipid analogs, including ceramide and sphingomyelin, has been reported ( 12,14,15 ). Importantly, acyl chain desaturation ( 17 ), deacylation/reacylation, and head group exchange ( 18 ) have been shown to occur in G. lamblia , indicating that the parasite can remodel the incorporated exogenous lipids to fulfi ll its own needs.…”

Section: Gene Expression Analysismentioning

confidence: 99%

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Glucosylceramide synthesis inhibition affects cell cycle progression, membrane trafficking, and stage differentiation in Giardia lamblia

Štefanić

Spycher

Morf

et al. 2010

Journal of Lipid Research

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Ceramide, a central molecule in the sphingolipid biosynthesis, plays a critical role as second messenger in cellular signaling that regulates antiproliferative processes, including apoptosis, cell differentiation, and cell cycle arrest in different cell types. Levels of ceramide, a highly bioactive molecule, must be tightly controlled by diverse, coordinated mechanisms, including ceramide degradation, phosphorylation, or sphingolipid metabolism. Glucosylceramide synthase (GCS), also defi ned as ceramide glucosyltransferase (CGT), metabolizes ceramide to glucosylceramide (GlcCer), a glycosylated form of ceramide that does not have antiproliferative activity. GCS plays a crucial role in cell survival after apoptotic stimuli, Abstract Synthesis of glucosylceramide via glucosylceramide synthase (GCS) is a crucial event in higher eukaryotes, both for the production of complex glycosphingolipids and for regulating cellular levels of ceramide, a potent antiproliferative second messenger. In this study, we explored the dependence of the early branching eukaryote Giardia lamblia on GCS activity. Biochemical analyses revealed that the parasite has a GCS located in endoplasmic reticulum (ER) membranes that is active in proliferating and encysting trophozoites. Pharmacological inhibition of GCS induced aberrant cell division, characterized by arrest of cytokinesis, incomplete cleavage furrow formation, and consequent block of replication. Importantly, we showed that increased ceramide levels were responsible for the cytokinesis arrest. In addition, GCS inhibition resulted in prominent ultrastructural abnormalities, including accumulation of cytosolic vesicles, enlarged lysosomes, and clathrin disorganization. Moreover, anterograde traffi cking of the encystations-specifi c protein CWP1 was severely compromised and resulted in inhibition of stage differentiation. Our results reveal novel aspects of lipid metabolism in G. lamblia and specifically highlight the vital role of GCS in regulating cell cycle progression, membrane traffi cking events, and stage differentiation in this parasite. In addition, we identifi ed ceramide as a potent bioactive molecule, underscoring the universal conservation of ceramide signaling in eukaryotes. -Štefanić,

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Section: Discussionmentioning

confidence: 99%

Section: Gene Expression Analysismentioning

confidence: 99%

Section: Gene Expression Analysismentioning

confidence: 99%

See 1 more Smart Citation

Glucosylceramide synthesis inhibition affects cell cycle progression, membrane trafficking, and stage differentiation in Giardia lamblia

Štefanić

Spycher

Morf

et al. 2010

Journal of Lipid Research

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“…Fatty acid desaturase activity, including desaturation of oleate to linoleate and linolenate, has been documented (75). Arachidonic acid is incorporated into neutral lipids, phospholipids, and a wide variety of cellular lipids (108), while palmitic acid, myristic acid, and oleic acid are transesterified primarily into phospholipids (28), including cellular phospholipids, (e.g., phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, and phosphatidylinositol) (149,154,228,313). Interesterification also occurs with incorporation of conjugated fatty acids into phosphatidylglycerol (108).…”

Section: Lipid Metabolismmentioning

confidence: 99%

Biology ofGiardia lamblia

2001

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Giardia lamblia is a common cause of diarrhea in humans and other mammals throughout the world. It can be distinguished from other Giardia species by light or electron microscopy. The two major genotypes of G. lamblia that infect humans are so different genetically and biologically that they may warrant separate species or subspecies designations. Trophozoites have nuclei and a well-developed cytoskeleton but lack mitochondria, peroxisomes, and the components of oxidative phosphorylation. They have an endomembrane system with at least some characteristics of the Golgi complex and encoplasmic reticulum, which becomes more extensive in encysting organisms. The primitive nature of the organelles and metabolism, as well as small-subunit rRNA phylogeny, has led to the proposal that Giardia spp. are among the most primitive eukaryotes. G. lamblia probably has a ploidy of 4 and a genome size of approximately 10 to 12 Mb divided among five chromosomes. Most genes have short 5′ and 3′ untranslated regions and promoter regions that are near the initiation codon. Trophozoites exhibit antigenic variation of an extensive repertoire of cysteine-rich variant-specific surface proteins. Expression is allele specific, and changes in expression from one vsp gene to another have not been associated with sequence alterations or gene rearrangements. The Giardia genome project promises to greatly increase our understanding of this interesting and enigmatic organism

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“…Some of the acquired lipids undergo remodeling by the head group and fatty acid exchange reactions. Fatty acids can undergo chain shortening or elongation before incorporation into the plasma membranes (10)(11)(12). Most recently, we have demonstrated that glucosylceramide transferase (GlcT1), an enzyme of the sphingolipid pathways, serves as a key regulator of encystation and viable cyst production by Giardia (13).…”

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confidence: 99%

The Assembly of GM1 Glycolipid- and Cholesterol-Enriched Raft-Like Membrane Microdomains Is Important for Giardial Encystation

et al. 2015

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Although encystation (or cyst formation) is an important step of the life cycle of Giardia, the cellular events that trigger encystation are poorly understood. Because membrane microdomains are involved in inducing growth and differentiation in many eukaryotes, we wondered if these raft-like domains are assembled by this parasite and participate in the encystation process. Since the GM1 ganglioside is a major constituent of mammalian lipid rafts (LRs) and known to react with cholera toxin B (CTXB), we used Alexa Fluor-conjugated CTXB and GM1 antibodies to detect giardial LRs. Raft-like structures in trophozoites are located in the plasma membranes and on the periphery of ventral discs. In cysts, however, they are localized in the membranes beneath the cyst wall. Nystatin and filipin III, two cholesterol-binding agents, and oseltamivir (Tamiflu), a viral neuraminidase inhibitor, disassembled the microdomains, as evidenced by reduced staining of trophozoites with CTXB and GM1 antibodies. GM1-and cholesterol-enriched LRs were isolated from Giardia by density gradient centrifugation and found to be sensitive to nystatin and oseltamivir. The involvement of LRs in encystation could be supported by the observation that raft inhibitors interrupted the biogenesis of encystation-specific vesicles and cyst production. Furthermore, culturing of trophozoites in dialyzed medium containing fetal bovine serum (which is low in cholesterol) reduced raft assembly and encystation, which could be rescued by adding cholesterol from the outside. Our results suggest that Giardia is able to form GM1-and cholesterol-enriched lipid rafts and these raft domains are important for encystation.

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Giardia lamblia:Incorporation of Free and Conjugated Fatty Acids into Glycerol-Based Phospholipids

Cited by 30 publications

References 33 publications

Glucosylceramide synthesis inhibition affects cell cycle progression, membrane trafficking, and stage differentiation in Giardia lamblia

Glucosylceramide synthesis inhibition affects cell cycle progression, membrane trafficking, and stage differentiation in Giardia lamblia

Biology ofGiardia lamblia

The Assembly of GM1 Glycolipid- and Cholesterol-Enriched Raft-Like Membrane Microdomains Is Important for Giardial Encystation

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