Publications by authors named "Samuel M Duncan"
Article Synopsis
- The bloodstream form of Trypanosoma brucei has large poly-N-acetyllactosamine (pNAL) chains on glycoproteins, which may be necessary for receptor-mediated endocytosis, and involves glycosyltransferases TbGT10 and TbGT8 in their biosynthesis.
- Researchers created TbGT10 and TbGT8 mutants to assess the impact of these enzymes on pNAL production, finding that although pNAL synthesis was reduced, it was not completely eliminated due to compensatory mechanisms from other glycosyltransferases.
- Interestingly, despite some glycoproteins being significantly affected by the pNAL deficiency, the mutants' transferrin receptor
Article Synopsis
- The biosynthesis of GPI-anchored proteins in Trypanosoma brucei requires fatty acid remodeling of GPI precursors before protein transfer in the endoplasmic reticulum.
- Researchers identified the gene Tb927.7.6110, which encodes a protein necessary for GPI-phospholipase A2 (GPI-PLA2) activity in the parasite's procyclic form.
- The absence of this gene in mutant cells led to reduced fatty acid remodeling and smaller GPI anchor sidechains, but this was restored by reintroducing Tb927.7.6110, indicating its critical role in the process.
Article Synopsis
- * These parasites retain some evolutionary components for N-glycosylation and GPI anchor synthesis but have evolved distinct glycosyltransferase families, particularly the GT67 family, which is linked to their life cycle and interactions with insect vectors.
- * A newly discovered GT11 family of fucosyltransferases, found only in the mitochondria of Trypanosoma brucei and Leishmania major, highlights the unique adaptations of these kinetoplastid parasites, with discussions on the
Article Synopsis
- The parasite Trypanosoma brucei has two forms—bloodstream (BSF) and procyclic (PCF)—that feature different carbohydrate structures linked to their survival and function, with at least 38 possible glycosyltransferase activities.* -
- Researchers investigated the uncharacterized glycosyltransferase gene TbGT10 and discovered it plays a key role in the formation of N-glycans in BSF and GPI anchors in PCF, implying it’s not essential for the parasite's survival despite impacting its fitness.* -
- Analysis showed that the absence of TbGT10 led to significant glycosylation defects, affecting how the parasite interacts with certain antibodies, highlighting its novel
Conditional gene deletion using dimerizable Cre recombinase (DiCre) is so far the best developed system for the phenotypic analysis of essential genes in Leishmania species. Here, we describe a protocol for the generation of a conditional gene deletion mutant and the subsequent inducible deletion of a target gene. Leishmania parasites are genetically modified to express two inactive Cre subunits (DiCre) and a single LoxP-flanked version of a target gene in a context where both endogenous copies of the gene have been deleted.
View Article and Find Full Text PDFLeishmania species are protozoan parasites whose remarkably plastic genome limits the establishment of effective genetic manipulation and leishmaniasis treatment. The strategies used by Leishmania to maintain its genome while allowing variability are not fully understood. Here, we used DiCre-mediated conditional gene deletion to show that HUS1, a component of the 9-1-1 (RAD9-RAD1-HUS1) complex, is essential and is required for a G2/M checkpoint.
View Article and Find Full Text PDFA DiCre recombinase-based system for inducible expression in Leishmania major.
Mol Biochem Parasitol
September 2017
Here we present the establishment of an inducible system based on the dimerizable Cre recombinase (DiCre) for controlled gene expression in the protozoan parasite Leishmania. Rapamycin-induced DiCre activation promoted efficient flipping and expression of gene products in a time and dose-dependent manner. The DiCre flipping activity induced the expression of target genes from both integrated and episomal contexts broadening the applicability of the system.
View Article and Find Full Text PDFRecent advances in Leishmania reverse genetics: Manipulating a manipulative parasite.
Mol Biochem Parasitol
September 2017
Article Synopsis
- * CRISPR/Cas9 approaches for large-scale gene knockout and tagging, as well as RNA manipulation methods like RNAi, are discussed to understand gene roles better.
- * Advancements in these molecular techniques aim to improve drug target validation and aid in developing new treatments for Leishmania-related diseases.
Leishmania mexicana has a large family of cyclin-dependent kinases (CDKs) that reflect the complex interplay between cell cycle and life cycle progression. Evidence from previous studies indicated that Cdc2-related kinase 3 (CRK3) in complex with the cyclin CYC6 is a functional homologue of the major cell cycle regulator CDK1, yet definitive genetic evidence for an essential role in parasite proliferation is lacking. To address this, we have implemented an inducible gene deletion system based on a dimerised Cre recombinase (diCre) to target CRK3 and elucidate its role in the cell cycle of L.
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