Eukaryotic Cell doi:10.1128/EC.00004-08
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.
The nascent parasitophorous vacuole membrane of E. cuniculi is formed by host cell lipids and contains pores which allow nutrient uptake
Karin Rönnebäumer,
Uwe Groß,
and
Wolfgang Bohne*
Institute of Medical Microbiology, University of Göttingen, Kreuzbergring 57, Göttingen D-37075; Germany
* To whom correspondence should be addressed. Email:
wbohne{at}gwdg.de.
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Abstract |
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Microsporidia are obligate intracellular pathogens, which enter host cells by the discharge of a hollow tube, through which the sporoplasma is extruded into the host cell. Since this invasion mechanism is very different from common entry strategies, the formation of the parasitophorous vacuole (PV) in Encephalitozoon species is likely to be distinct from known principles. We investigated the origin of the nascent E. cuniculi PV membrane with the aid of fluorescent lipid probes. When BODIPY-500/510-C12-HPC labeled spores were used for infection, the emerging PV membrane was unlabeled, suggesting that sporoplasma-derived lipids do not significantly contribute to the formation of the PV membrane. In contrast, when raft and nonraft microdomains of the host cell plasma membrane were selectively labeled with DiIC16 and SPEEDY DiO, both tracers were detectable in the nascent PV membrane shortly after infection, indicating that the bulk lipids of the PV membrane are host cell derived. Time lapse fluorescence microscopy revealed that the formation of the PV membrane is a fast event (< 1.3 sec), which occurred simultaneously with the extrusion of the sporoplasma. The portion of the discharged tube which is in contact with the host cell was found to be coated with labeled host cell lipids, which might be an indication for a plasma membrane invagination at the contact site. To investigate the presence of pores in the E. cuniculi PV membrane, we microinjected fluorescent dyes of different sizes into infected host cells. A 0.5 kDa dextran as well as 0.8-1.1 kDa peptides could rapidly enter the PV while a 10 kDa dextran was stably excluded from the PV lumen, indicating that the PV membrane possesses pores with an exclusion size of <10 kDa, which should allow metabolite exchange.
