SummaryAs Giardia lamblia is unable to synthesize cholesterol de novo, this steroid might be obtained from the host's intestinal milieu by endocytosis of lipoproteins. In this work, we identified a putative Giardia lamblia low-density lipoprotein receptorrelated proteins (GlLRP), a type I membrane protein, which shares the substrate N-terminal binding domain and a FXNPXY-type endocytic motif with human LRPs. Expression of tagged GlLRP showed that it was localized predominantly in the endoplasmic reticulum, lysosomal-like peripheral vacuoles and plasma membrane. However, the FXNPXYdeleted GlLRP was retained at the plasma membrane suggesting that it is abnormally transported and processed. The low-density lipoprotein and chylomicrons interacted with GlLRP, with this interaction being necessary for lipoprotein internalization and cell proliferation. Finally, we show that GlLRP binds directly to the medium subunit of Giardia adaptor protein 2, indicating that receptor-mediated internalization occurs through an adaptin mechanism.
In Giardia, lysosome-like peripheral vacuoles (PVs) need to specifically coordinate their endosomal and lysosomal functions to be able to successfully perform endocytosis, protein degradation and protein delivery, but how cargo, ligands and molecular components generate specific routes to the PVs remains poorly understood. Recently, we found that delivering membrane Cathepsin C and the soluble acid phosphatase (AcPh) to the PVs is adaptin (AP1)-dependent. However, the receptor that links AcPh and AP1 was never described. We have studied protein-binding to AcPh by using H6-tagged AcPh, and found that a membrane protein interacted with AcPh. This protein, named GlVps (for Giardia lamblia Vacuolar protein sorting), mainly localized to the ER-nuclear envelope and in some PVs, probably functioning as the sorting receptor for AcPh. The tyrosine-binding motif found in the C-terminal cytoplasmic tail domain of GlVps was essential for its exit from the endoplasmic reticulum and transport to the vacuoles, with this motif being necessary for the interaction with the medium subunit of AP1. Thus, the mechanism by which soluble proteins, such as AcPh, reach the peripheral vacuoles in Giardia appears to be very similar to the mechanism of lysosomal protein-sorting in more evolved eukaryotic cells.
Aims: To study genomic and phenotypic changes in wine yeasts produced in short time periods analysing yeast strains possibly derived from commercial strains recently dispersed.
Methods and Results: We conducted a genomic and phenotypic comparison between the commercial yeast strain EC1118 and two novel strains (LV CB and L‐957) isolated from different wine areas industrially intervened <20 years ago. Molecular analysis by amplified fragment length polymorphism (AFLP) and RAPD‐PCR was not able to distinguish between these strains. However, comparative genomic hybridization (aCGH) showed discrete DNA gains and losses that allowed unequivocal identification of the strains. Furthermore, analysis of aCGH data supports the hypothesis that strains LV CB and L‐957 are derivatives from strain EC1118. Finally, scarce phenotypic differences in physiological and metabolic parameters were found among the strains.
Conclusion: The wine yeasts have a very dynamic genome that accumulates changes in short time periods. These changes permit the unique genomic identification of the strains.
Significance and Impact of the Study: This study permits the evaluation of microevolutive events in wine yeasts and its relationship with the phenotype in this species.
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