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A Genetic Screen to Isolate Toxoplasma gondii Host-cell Egress Mutants
Published on: February 8, 2012
Molecular partitioning during host cell penetration by Toxoplasma gondii
Audra J Charron1, L David Sibley
1Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, MO 63110, USA.
Abstract:
During invasion by Toxoplasma gondii, host cell transmembrane proteins are excluded from the forming parasitophorous vacuole membrane (PVM) by the tight apposition of host and parasite cellular membranes. Previous studies suggested that the basis for the selective partitioning of membrane constituents may be a preference for membrane microdomains, and this hypothesis was herein tested. The partitioning of a diverse group of molecular reporters for raft and nonraft membrane subdomains was monitored during parasite invasion by time-lapse video or confocal microscopy. Unexpectedly, both raft and nonraft lipid probes, as well as both raft and nonraft cytosolic leaflet proteins, flowed unhindered past the host-parasite junction into the PVM. Moreover, neither a raft-associated type 1 transmembrane protein nor its raft-dissociated counterpart accessed the PVM, while a multispanning membrane raft protein readily did so. Considered together with previous data, these studies demonstrate that selective partitioning at the host-parasite interface is a highly complex process, in which raft association favors, but is neither necessary nor sufficient for, inclusion into the T. gondii PVM.
Insights
During Toxoplasma gondii invasion, host proteins are excluded from the parasitophorous vacuole membrane (PVM). Membrane microdomain preference is not the sole factor; raft association influences, but doesn't guarantee, PVM inclusion.
Area of Science:
- Cell Biology
- Parasitology
- Membrane Biophysics
Background:
- Host cell proteins are typically excluded from the parasitophorous vacuole membrane (PVM) during Toxoplasma gondii invasion.
- Previous research suggested membrane microdomains might dictate selective partitioning of host membrane components into the PVM.
Purpose of the Study:
- To investigate the role of membrane microdomains (rafts and non-rafts) in the selective inclusion of host proteins into the PVM during T. gondii invasion.
- To test the hypothesis that preferential association with membrane microdomains drives protein partitioning into the PVM.
Main Methods:
- Utilized time-lapse video and confocal microscopy to monitor molecular reporters.
- Assessed partitioning of various lipid probes and cytosolic leaflet proteins representing raft and non-raft domains.
- Examined the behavior of specific transmembrane proteins, including raft-associated and raft-dissociated types, and a multispanning raft protein.
Main Results:
- Both raft and non-raft lipid probes, along with raft and non-raft cytosolic proteins, freely entered the PVM.
- Neither a raft-associated type 1 transmembrane protein nor its raft-dissociated counterpart were found in the PVM.
- A multispanning membrane raft protein was readily incorporated into the PVM.
Conclusions:
- Selective partitioning of host membrane constituents into the T. gondii PVM is a complex process.
- Raft association is not the sole determinant; it favors, but is neither necessary nor sufficient for, PVM inclusion.
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