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Differential Translocation of Host Cellular Materials into the Chlamydia trachomatis Inclusion Lumen during Chemical Fixation.

Kokes M, Valdivia RH - PLoS ONE (2015)

Bottom Line: However, we see little evidence of intraluminal localization of these organelles in live inclusions.These intra-inclusion ER elements resist a variety of post-fixation manipulations and are detectable via immunofluorescence microscopy.Finally, we find similar structures within the pathogenic vacuole of Coxiella burnetti infected cells, suggesting that fixation-induced translocation of cellular materials may occur into the vacuole of a range of intracellular pathogens.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology and Center for the Genomics of Microbial Systems, Duke University Medical Center, Durham, North Carolina, United States of America.

ABSTRACT
Chlamydia trachomatis manipulates host cellular pathways to ensure its proliferation and survival. Translocation of host materials into the pathogenic vacuole (termed 'inclusion') may facilitate nutrient acquisition and various organelles have been observed within the inclusion, including lipid droplets, peroxisomes, multivesicular body components, and membranes of the endoplasmic reticulum (ER). However, few of these processes have been documented in living cells. Here, we survey the localization of a broad panel of subcellular elements and find ER, mitochondria, and inclusion membranes within the inclusion lumen of fixed cells. However, we see little evidence of intraluminal localization of these organelles in live inclusions. Using time-lapse video microscopy we document ER marker translocation into the inclusion lumen during chemical fixation. These intra-inclusion ER elements resist a variety of post-fixation manipulations and are detectable via immunofluorescence microscopy. We speculate that the localization of a subset of organelles may be exaggerated during fixation. Finally, we find similar structures within the pathogenic vacuole of Coxiella burnetti infected cells, suggesting that fixation-induced translocation of cellular materials may occur into the vacuole of a range of intracellular pathogens.

No MeSH data available.


Related in: MedlinePlus

ER-RFP within the pathogenic vacuole of Coxiella burnettii.HeLa cells were infected with Coxiella burnettii, transfected with ER-RFP, and fixed at 52 hpi. Note the presence of ER-RFP structures with in the lumen of the pathogenic vacuole (cyan arrowheads) similar to those within Chlamydia trachomatis inclusions. N indicates the nucleus and a dashed line outlines a region of higher magnification. Scale bar represents 5 μm.
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pone.0139153.g005: ER-RFP within the pathogenic vacuole of Coxiella burnettii.HeLa cells were infected with Coxiella burnettii, transfected with ER-RFP, and fixed at 52 hpi. Note the presence of ER-RFP structures with in the lumen of the pathogenic vacuole (cyan arrowheads) similar to those within Chlamydia trachomatis inclusions. N indicates the nucleus and a dashed line outlines a region of higher magnification. Scale bar represents 5 μm.

Mentions: The Chlamydia inclusion is an unusually spacious organelle and the luminal space is largely devoid of electron-dense material compared to the host cytoplasm as assessed by transmission electron microscopy [39]. We speculated that this spacious nature may be conducive to fixation-induced translocation of materials that might otherwise not occur elsewhere in the cell. To assess this, we asked whether another similarly spacious pathogenic vacuole occupied by Coxiella burnettii (reviewed in [40]) would display similar internalized structures. When we transfected Coxiella burnettii-infected cells with ER-RFP and fixed after 52 hr of infection, we found ER-RFP structures within the lumen of the pathogenic vacuole with similar appearance and frequency (90%) as those found within Chlamydia inclusions (Fig 5).


Differential Translocation of Host Cellular Materials into the Chlamydia trachomatis Inclusion Lumen during Chemical Fixation.

Kokes M, Valdivia RH - PLoS ONE (2015)

ER-RFP within the pathogenic vacuole of Coxiella burnettii.HeLa cells were infected with Coxiella burnettii, transfected with ER-RFP, and fixed at 52 hpi. Note the presence of ER-RFP structures with in the lumen of the pathogenic vacuole (cyan arrowheads) similar to those within Chlamydia trachomatis inclusions. N indicates the nucleus and a dashed line outlines a region of higher magnification. Scale bar represents 5 μm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4591358&req=5

pone.0139153.g005: ER-RFP within the pathogenic vacuole of Coxiella burnettii.HeLa cells were infected with Coxiella burnettii, transfected with ER-RFP, and fixed at 52 hpi. Note the presence of ER-RFP structures with in the lumen of the pathogenic vacuole (cyan arrowheads) similar to those within Chlamydia trachomatis inclusions. N indicates the nucleus and a dashed line outlines a region of higher magnification. Scale bar represents 5 μm.
Mentions: The Chlamydia inclusion is an unusually spacious organelle and the luminal space is largely devoid of electron-dense material compared to the host cytoplasm as assessed by transmission electron microscopy [39]. We speculated that this spacious nature may be conducive to fixation-induced translocation of materials that might otherwise not occur elsewhere in the cell. To assess this, we asked whether another similarly spacious pathogenic vacuole occupied by Coxiella burnettii (reviewed in [40]) would display similar internalized structures. When we transfected Coxiella burnettii-infected cells with ER-RFP and fixed after 52 hr of infection, we found ER-RFP structures within the lumen of the pathogenic vacuole with similar appearance and frequency (90%) as those found within Chlamydia inclusions (Fig 5).

Bottom Line: However, we see little evidence of intraluminal localization of these organelles in live inclusions.These intra-inclusion ER elements resist a variety of post-fixation manipulations and are detectable via immunofluorescence microscopy.Finally, we find similar structures within the pathogenic vacuole of Coxiella burnetti infected cells, suggesting that fixation-induced translocation of cellular materials may occur into the vacuole of a range of intracellular pathogens.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology and Center for the Genomics of Microbial Systems, Duke University Medical Center, Durham, North Carolina, United States of America.

ABSTRACT
Chlamydia trachomatis manipulates host cellular pathways to ensure its proliferation and survival. Translocation of host materials into the pathogenic vacuole (termed 'inclusion') may facilitate nutrient acquisition and various organelles have been observed within the inclusion, including lipid droplets, peroxisomes, multivesicular body components, and membranes of the endoplasmic reticulum (ER). However, few of these processes have been documented in living cells. Here, we survey the localization of a broad panel of subcellular elements and find ER, mitochondria, and inclusion membranes within the inclusion lumen of fixed cells. However, we see little evidence of intraluminal localization of these organelles in live inclusions. Using time-lapse video microscopy we document ER marker translocation into the inclusion lumen during chemical fixation. These intra-inclusion ER elements resist a variety of post-fixation manipulations and are detectable via immunofluorescence microscopy. We speculate that the localization of a subset of organelles may be exaggerated during fixation. Finally, we find similar structures within the pathogenic vacuole of Coxiella burnetti infected cells, suggesting that fixation-induced translocation of cellular materials may occur into the vacuole of a range of intracellular pathogens.

No MeSH data available.


Related in: MedlinePlus