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Sequestration of host metabolism by an intracellular pathogen.

Gehre L, Gorgette O, Perrinet S, Prevost MC, Ducatez M, Giebel AM, Nelson DE, Ball SG, Subtil A - Elife (2016)

Bottom Line: We provide evidence that bacterial glycogen metabolism enzymes are secreted into the vacuole lumen through type 3 secretion.Our data bring strong support to the following scenario: bacteria co-opt the host transporter SLC35D2 to import UDP-glucose into the vacuole, where it serves as substrate for de novo glycogen synthesis, through a remarkable adaptation of the bacterial glycogen synthase.Based on these findings we propose that parasitophorous vacuoles not only offer protection but also provide a microorganism-controlled metabolically active compartment essential for redirecting host resources to the pathogens.

View Article: PubMed Central - PubMed

Affiliation: Unité de Biologie cellulaire de l'infection microbienne, Institut Pasteur, Paris, France.

ABSTRACT
For intracellular pathogens, residence in a vacuole provides a shelter against cytosolic host defense to the cost of limited access to nutrients. The human pathogen Chlamydia trachomatis grows in a glycogen-rich vacuole. How this large polymer accumulates there is unknown. We reveal that host glycogen stores shift to the vacuole through two pathways: bulk uptake from the cytoplasmic pool, and de novo synthesis. We provide evidence that bacterial glycogen metabolism enzymes are secreted into the vacuole lumen through type 3 secretion. Our data bring strong support to the following scenario: bacteria co-opt the host transporter SLC35D2 to import UDP-glucose into the vacuole, where it serves as substrate for de novo glycogen synthesis, through a remarkable adaptation of the bacterial glycogen synthase. Based on these findings we propose that parasitophorous vacuoles not only offer protection but also provide a microorganism-controlled metabolically active compartment essential for redirecting host resources to the pathogens.

No MeSH data available.


Related in: MedlinePlus

Flag-GlgA is imported into the inclusion lumen and enhances luminal glycogen accumulation.(A) HeLa cells were transfected (top right) or not (top left) with Flag-GlgA before infection, and fixed 24 hpi. PAS staining revealed an increase of intraluminal glycogen (arrowheads) in Flag-GlgA expressing cells. C. trachomatis has a 8 kb plasmid, and its loss is associated with decreased GlgA expression and impaired glycogen accumulation (Carlson et al., 2008) (bottom left). Remarkably, transfection of Flag-GlgA restored luminal glycogen accumulation in cells infected with the plasmid-less strain LGV 25667R (bottom right). (B) Immunofluorescence on Flag-GlgA transfected cells infected with the wild-type LGV strain. DNA is stained in blue, Flag-GlgA in green and the inclusion membrane in red (anti-Cap1). Flag-GlgA is abundant in the host cytoplasm and the inclusion lumen (see also xz (top) and yz (right) projections). Scale bars: 10 µm.DOI:http://dx.doi.org/10.7554/eLife.12552.016
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fig5s1: Flag-GlgA is imported into the inclusion lumen and enhances luminal glycogen accumulation.(A) HeLa cells were transfected (top right) or not (top left) with Flag-GlgA before infection, and fixed 24 hpi. PAS staining revealed an increase of intraluminal glycogen (arrowheads) in Flag-GlgA expressing cells. C. trachomatis has a 8 kb plasmid, and its loss is associated with decreased GlgA expression and impaired glycogen accumulation (Carlson et al., 2008) (bottom left). Remarkably, transfection of Flag-GlgA restored luminal glycogen accumulation in cells infected with the plasmid-less strain LGV 25667R (bottom right). (B) Immunofluorescence on Flag-GlgA transfected cells infected with the wild-type LGV strain. DNA is stained in blue, Flag-GlgA in green and the inclusion membrane in red (anti-Cap1). Flag-GlgA is abundant in the host cytoplasm and the inclusion lumen (see also xz (top) and yz (right) projections). Scale bars: 10 µm.DOI:http://dx.doi.org/10.7554/eLife.12552.016

Mentions: Our data strongly support the hypothesis that chlamydial glycogen synthase (GlgA) initiates de novo glycogen synthesis in the inclusion lumen. This raises the question of its substrate specificity. A sharp distinction between prokaryote and eukaryote glycogen synthases is that, almost without exception, bacterial glycogen synthases function on ADP-Glc while eukaryotic glycogen synthases use UDP-Glc as substrate. We reasoned that GlgA secretion into the host cytoplasm, which contains no ADP-Glc, might point to an unusual substrate specificity for this enzyme. Transfection of cells with flag-tagged chlamydial GlgA led to massive glycogen accumulation, proving that indeed C. trachomatis GlgA is functional on UDP-Glc (Figure 5A). Interestingly, when transfected cells were subsequently infected, an increase in glycogen accumulation in the inclusion lumen was observed, and Flag-GlgA was detected in the bacterial compartment (Figure 5—figure supplement 1). High intraluminal glycogen content upon ectopic GlgA expression was also observed when cells were infected with a strain devoid of the natural plasmid of C. trachomatis (Figure 5—figure supplement 1), and Flag-GlgA was observed in the inclusion lumen also in that case (not shown). Plasmid-loss is associated with decreased GlgA expression and impaired glycogen accumulation (Carlson et al., 2008). In non-transfected cells the polysaccharide was still detected in EBs of the plasmid-less strain and, to a highly reduced level, in the inclusion lumen (Figure 5—figure supplement 2). Glycogen recovery upon GlgA transfection indicates that the low level of expression of GlgA in the plasmid-less strain accounts for the defect in glycogen storage. It is quite remarkable that a protein expressed by the host can compensate for a bacterial deficiency.10.7554/eLife.12552.015Figure 5.Host UDP-Glc is the substrate for intraluminal glycogen synthesis.


Sequestration of host metabolism by an intracellular pathogen.

Gehre L, Gorgette O, Perrinet S, Prevost MC, Ducatez M, Giebel AM, Nelson DE, Ball SG, Subtil A - Elife (2016)

Flag-GlgA is imported into the inclusion lumen and enhances luminal glycogen accumulation.(A) HeLa cells were transfected (top right) or not (top left) with Flag-GlgA before infection, and fixed 24 hpi. PAS staining revealed an increase of intraluminal glycogen (arrowheads) in Flag-GlgA expressing cells. C. trachomatis has a 8 kb plasmid, and its loss is associated with decreased GlgA expression and impaired glycogen accumulation (Carlson et al., 2008) (bottom left). Remarkably, transfection of Flag-GlgA restored luminal glycogen accumulation in cells infected with the plasmid-less strain LGV 25667R (bottom right). (B) Immunofluorescence on Flag-GlgA transfected cells infected with the wild-type LGV strain. DNA is stained in blue, Flag-GlgA in green and the inclusion membrane in red (anti-Cap1). Flag-GlgA is abundant in the host cytoplasm and the inclusion lumen (see also xz (top) and yz (right) projections). Scale bars: 10 µm.DOI:http://dx.doi.org/10.7554/eLife.12552.016
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Related In: Results  -  Collection

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fig5s1: Flag-GlgA is imported into the inclusion lumen and enhances luminal glycogen accumulation.(A) HeLa cells were transfected (top right) or not (top left) with Flag-GlgA before infection, and fixed 24 hpi. PAS staining revealed an increase of intraluminal glycogen (arrowheads) in Flag-GlgA expressing cells. C. trachomatis has a 8 kb plasmid, and its loss is associated with decreased GlgA expression and impaired glycogen accumulation (Carlson et al., 2008) (bottom left). Remarkably, transfection of Flag-GlgA restored luminal glycogen accumulation in cells infected with the plasmid-less strain LGV 25667R (bottom right). (B) Immunofluorescence on Flag-GlgA transfected cells infected with the wild-type LGV strain. DNA is stained in blue, Flag-GlgA in green and the inclusion membrane in red (anti-Cap1). Flag-GlgA is abundant in the host cytoplasm and the inclusion lumen (see also xz (top) and yz (right) projections). Scale bars: 10 µm.DOI:http://dx.doi.org/10.7554/eLife.12552.016
Mentions: Our data strongly support the hypothesis that chlamydial glycogen synthase (GlgA) initiates de novo glycogen synthesis in the inclusion lumen. This raises the question of its substrate specificity. A sharp distinction between prokaryote and eukaryote glycogen synthases is that, almost without exception, bacterial glycogen synthases function on ADP-Glc while eukaryotic glycogen synthases use UDP-Glc as substrate. We reasoned that GlgA secretion into the host cytoplasm, which contains no ADP-Glc, might point to an unusual substrate specificity for this enzyme. Transfection of cells with flag-tagged chlamydial GlgA led to massive glycogen accumulation, proving that indeed C. trachomatis GlgA is functional on UDP-Glc (Figure 5A). Interestingly, when transfected cells were subsequently infected, an increase in glycogen accumulation in the inclusion lumen was observed, and Flag-GlgA was detected in the bacterial compartment (Figure 5—figure supplement 1). High intraluminal glycogen content upon ectopic GlgA expression was also observed when cells were infected with a strain devoid of the natural plasmid of C. trachomatis (Figure 5—figure supplement 1), and Flag-GlgA was observed in the inclusion lumen also in that case (not shown). Plasmid-loss is associated with decreased GlgA expression and impaired glycogen accumulation (Carlson et al., 2008). In non-transfected cells the polysaccharide was still detected in EBs of the plasmid-less strain and, to a highly reduced level, in the inclusion lumen (Figure 5—figure supplement 2). Glycogen recovery upon GlgA transfection indicates that the low level of expression of GlgA in the plasmid-less strain accounts for the defect in glycogen storage. It is quite remarkable that a protein expressed by the host can compensate for a bacterial deficiency.10.7554/eLife.12552.015Figure 5.Host UDP-Glc is the substrate for intraluminal glycogen synthesis.

Bottom Line: We provide evidence that bacterial glycogen metabolism enzymes are secreted into the vacuole lumen through type 3 secretion.Our data bring strong support to the following scenario: bacteria co-opt the host transporter SLC35D2 to import UDP-glucose into the vacuole, where it serves as substrate for de novo glycogen synthesis, through a remarkable adaptation of the bacterial glycogen synthase.Based on these findings we propose that parasitophorous vacuoles not only offer protection but also provide a microorganism-controlled metabolically active compartment essential for redirecting host resources to the pathogens.

View Article: PubMed Central - PubMed

Affiliation: Unité de Biologie cellulaire de l'infection microbienne, Institut Pasteur, Paris, France.

ABSTRACT
For intracellular pathogens, residence in a vacuole provides a shelter against cytosolic host defense to the cost of limited access to nutrients. The human pathogen Chlamydia trachomatis grows in a glycogen-rich vacuole. How this large polymer accumulates there is unknown. We reveal that host glycogen stores shift to the vacuole through two pathways: bulk uptake from the cytoplasmic pool, and de novo synthesis. We provide evidence that bacterial glycogen metabolism enzymes are secreted into the vacuole lumen through type 3 secretion. Our data bring strong support to the following scenario: bacteria co-opt the host transporter SLC35D2 to import UDP-glucose into the vacuole, where it serves as substrate for de novo glycogen synthesis, through a remarkable adaptation of the bacterial glycogen synthase. Based on these findings we propose that parasitophorous vacuoles not only offer protection but also provide a microorganism-controlled metabolically active compartment essential for redirecting host resources to the pathogens.

No MeSH data available.


Related in: MedlinePlus