Dataset Information

Stage-specific, morphological and molecular markers of encystation in Giardia lamblia

ABSTRACT: Differentiation into environmentally resistant cysts is required for transmission of the ubiquitous intestinal parasite Giardia lamblia. Encystation in Giardia requires the production, processing and transport of Cyst Wall Proteins (CWPs) in stage induced Golgi-like Encystation Specific Vesicles (ESVs). While progress of this trafficking pathway can be followed by tracking CWPs, there is no recognized system to distinguish stages of this process that takes varied lengths of time to complete depending on the encystation method used. Here we propose a staging system for encysting Giardia based on the morphology of CWP1-stained ESVs. The molecular distinctiveness of ESVs at these stages were demonstrated by following Giardia Rabs through encystation. We previously demonstrated that Giardia’s sole Rho family GTPase, GlRac, associates with ESVs and has a role in regulating CWP1 maturation and secretion. As a proof of principle, we delineated the relationship among GlRac and ESV stages and through proteomic studies identified putative interactors of GlRac that could be used as additional ESV Stage-specific markers. We propose that this staging system provides a common descriptor of ESV maturation regardless of the source of encysting cells and the identified set of molecular markers for different ESV stages will be a powerful tool for characterizing trafficking mutants that impair typical ESV maturation and morphology.

INSTRUMENT(S): LTQ

ORGANISM(S): Giardia Lamblia Atcc 50803

TISSUE(S): Permanent Cell Line Cell, Cell Culture

DISEASE(S): Giardiasis

SUBMITTER: Richard Johnson

LAB HEAD: Alexander Paradez

PROVIDER: PXD024944 | Pride | 2021-07-10

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	Rac8henc_VP_2017_0131_RJ_racVwt__8h_enc_2nd_and_3rd_.xlsx	Xlsx
	Rac_8h_encyst_and_troph_vs_WT_11032016__troph_3rd_and_8h_enc_1st_.xlsx	Xlsx
	Rac_v_wt_troph_2016_0809__troph_2nd_.xlsx	Xlsx
	RacvsWT_with_fisher_exact_test__troph_1st__FisherExact.xlsx	Xlsx
	VP_2014_0513_RJ_07_wt.raw	Raw

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Publications

Staging Encystation Progression in Giardia lamblia Using Encystation-Specific Vesicle Morphology and Associating Molecular Markers.

Thomas Elizabeth B EB Sutanto Renaldo R Johnson Richard S RS Shih Han-Wei HW Alas Germain C M GCM Krtková Jana J MacCoss Michael J MJ Paredez Alexander R AR

Frontiers in cell and developmental biology 20210427

Differentiation into environmentally resistant cysts is required for transmission of the ubiquitous intestinal parasite Giardia lamblia. Encystation in Giardia requires the production, processing and transport of Cyst Wall Proteins (CWPs) in developmentally induced, Golgi-like, Encystation Specific Vesicles (ESVs). Progress through this trafficking pathway can be followed by tracking CWP localization over time. However, there is no recognized system to distinguish the advancing sta ...[more]

PMID: 33987184

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Project description:Background: Cyst formation in parasitic protist Giardia duodenalis is critical to its transmission. Proteomic data is essential for the system-level understanding of the molecular mechanisms managing stage transition from a flagellated, binucleate trophozoite to a tetra-nucleate cyst. The current encystation proteome quantifies 16% of predicted protein coding genes and lacks complete coverage of the developmental transition. Given recent gains in mass spectrometry instrumentation, we have generated significantly improved proteomes for Giardia across encystation, providing framework for transcriptome-proteome correlation drawing insights into molecular networks modulating encystation in Giardia. Results: We have reproducibly identified 3,796 (64.6% of Giardia proteins) and quantified 3,402 proteins (56.12 % of Giardia proteins), tripling the currently published encystation proteome coverage. This includes timepoints from standard trophozoite growth (TY), during low-bile encystation priming (LB), mid-encystation (EC), and enriched, mature cysts (C), making this the first proteome study across the entire encystation process. This is also the first proteome analysis of trophozoites during low bile priming prior to in vitro induction of encystation, and analysis of mature cyst proteomes. Almost 28% (1,102 proteins) of the proteome is differentially changed during encystation including within proteasomal machinery, metabolic pathways and lipid metabolism, indicating a multimodal regulation in encystation. To further understand novel observations in lipid metabolism, we used hidden markov model (HMM) based domain prediction strategies and i-Tasser based structural modelling to provide evidence for the presence of three families of lipid transporters (LTP’s) StARkin (steroidogenic acute regulatory protein–related lipid transfer), ORP/Osh (Oxysterol Binding protein related protein), GLTP (Glycosphingolipid transfer protein) in Giardia. Finally, we performed functional interaction studies using in silico docking of newly identified LTP’s with corresponding lipid ligands to validate their interaction. This identified a shift in lipid species dependency in Giardia as the parasite progress towards cyst formation.

Dataset Information

Stage-specific, morphological and molecular markers of encystation in Giardia lamblia

Dataset's files

Publications

Staging Encystation Progression in <i>Giardia lamblia</i> Using Encystation-Specific Vesicle Morphology and Associating Molecular Markers.

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