Glucosylceramide Transferase Activity Is Critical for Encystation and Viable Cyst Production by an Intestinal Protozoan, Giardia lamblia

• Published in: The Journal of biological chemistry 2013-06, Vol.288 (23), p.16747-16760
• Main Authors: Mendez, Tavis L., De Chatterjee, Atasi, Duarte, Trevor T., Gazos-Lopes, Felipe, Robles-Martinez, Leobarda, Roy, Debarshi, Sun, Jianjun, Maldonado, Rosa A., Roychowdhury, Sukla, Almeida, Igor C., Das, Siddhartha
• Format: Article
• Language: English
• Subjects: Giardia; Giardia lamblia - enzymology; Giardia lamblia - genetics; Giardia lamblia - growth & development; Glucosylceramide; Glycolipids; Glycosyltransferases - genetics; Glycosyltransferases - metabolism; Humans; Lipid Metabolism; Lipids; Parasite; Parasite Metabolism; Protozoan Proteins - genetics; Protozoan Proteins - metabolism; Sphingolipid; Sphingolipids - biosynthesis; Sphingolipids - genetics; Transferase
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M112.438416

The production of viable cysts by Giardia is essential for its survival in the environment and for spreading the infection via contaminated food and water. The hallmark of cyst production (also known as encystation) is the biogenesis of encystation-specific vesicles (ESVs) that transport cyst wall proteins to the plasma membrane of the trophozoite before laying down the protective cyst wall. However, the molecules that regulate ESV biogenesis and maintain cyst viability have never before been identified. Here, we report that giardial glucosylceramide transferase-1 (gGlcT1), an enzyme of sphingolipid biosynthesis, plays a key role in ESV biogenesis and maintaining cyst viability. We find that overexpression of this enzyme induced the formation of aggregated/enlarged ESVs and generated clustered cysts with reduced viability. The silencing of gGlcT1 synthesis by antisense morpholino oligonucleotide abolished ESV production and generated mostly nonviable cysts. Interestingly, when gGlcT1-overexpressed Giardia was transfected with anti-gGlcT1 morpholino, the enzyme activity, vesicle biogenesis, and cyst viability returned to normal, suggesting that the regulated expression of gGlcT1 is important for encystation and viable cyst production. Furthermore, the overexpression of gGlcT1 increased the influx of membrane lipids and fatty acids without altering the fluidity of plasma membranes, indicating that the expression of gGlcT1 activity is linked to lipid internalization and maintaining the overall lipid balance in this parasite. Taken together, our results suggest that gGlcT1 is a key player of ESV biogenesis and cyst viability and therefore could be targeted for developing new anti-giardial therapies. Background: The production of viable cysts by Giardia is essential for transmitting the infection via contaminated food and water. Results: Overexpression and knockdown of glucosylceramide transferase activity affect encystation, cyst viability, and overall lipid balance in Giardia. Conclusion: Regulated expression of glucosylceramide transferase is linked to encystation and cyst production. Significance: Glucosylceramide synthesis could be targeted for developing novel anti-giardial therapy.