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Cellular prion protein promotes Brucella infection into macrophages.

Watarai M, Kim S, Erdenebaatar J, Makino S, Horiuchi M, Shirahata T, Sakaguchi S, Katamine S - J. Exp. Med. (2003)

Bottom Line: Hsp60 reacted strongly with serum from human brucellosis patients and was exposed on the bacterial surface via a VirB complex-associated process.Under in vitro and in vivo conditions, Hsp60 of B. abortus bound to PrPC.These results indicate that signal transduction induced by the interaction between bacterial Hsp60 and PrPC on macrophages contributes to the establishment of B. abortus infection.

View Article: PubMed Central - PubMed

Affiliation: Department of Applied Veterinary Science, Obihiro University of Agriculture and Veterinary Medicine, Inada-cho, Obihiro-shi, Hokkaido 080-8555, Japan. watarai@obihiro.ac.jp

ABSTRACT
The products of the Brucella abortus virB gene locus, which are highly similar to conjugative DNA transfer system, enable the bacterium to replicate within macrophage vacuoles. The replicative phagosome is thought to be established by the interaction of a substrate of the VirB complex with macrophages, although the substrate and its host cellular target have not yet been identified. We report here that Hsp60, a member of the GroEL family of chaperonins, of B. abortus is capable of interacting directly or indirectly with cellular prion protein (PrPC) on host cells. Aggregation of PrPC tail-like formation was observed during bacterial swimming internalization into macrophages and PrPC was selectively incorporated into macropinosomes containing B. abortus. Hsp60 reacted strongly with serum from human brucellosis patients and was exposed on the bacterial surface via a VirB complex-associated process. Under in vitro and in vivo conditions, Hsp60 of B. abortus bound to PrPC. Hsp60 of B. abortus, expressed on the surface of Lactococcus lactis, promoted the aggregation of PrPC but not PrPC tail formation on macrophages. The PrPC deficiency prevented swimming internalization and intracellular replication of B. abortus, with the result that phagosomes bearing the bacteria were targeted into the endocytic network. These results indicate that signal transduction induced by the interaction between bacterial Hsp60 and PrPC on macrophages contributes to the establishment of B. abortus infection.

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Tail formation of GPI-anchored proteins on the site of swimming internalization. Bone marrow–derived macrophages were incubated with B. abortus for 5 or 15 min, and GPI-anchored proteins were localized by immunofluorescence as described in Materials and Methods. Merged images of the GFP (green) and TRITC (red) channels up and down with phase contrast images of the same field are shown. Cells were probed with aerolysin for GPI-anchored proteins and with anti-CD48. White arrows point to bacteria and blue arrow points to tail-like aggregation of GPI-anchored proteins.
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fig1: Tail formation of GPI-anchored proteins on the site of swimming internalization. Bone marrow–derived macrophages were incubated with B. abortus for 5 or 15 min, and GPI-anchored proteins were localized by immunofluorescence as described in Materials and Methods. Merged images of the GFP (green) and TRITC (red) channels up and down with phase contrast images of the same field are shown. Cells were probed with aerolysin for GPI-anchored proteins and with anti-CD48. White arrows point to bacteria and blue arrow points to tail-like aggregation of GPI-anchored proteins.

Mentions: Our previous results showed that GPI-anchored proteins were selectively incorporated into macropinosomes containing B. abortus (Fig. 1; reference 6). To investigate further the membrane sorting process, the distribution of GPI-anchored proteins during swimming internalization of B. abortus was analyzed. Aerolysin from Aeromonas hydrophila, which binds to the GPI moiety of GPI-anchored proteins on the cell surface (26), was used as probe for the detection of GPI-anchored proteins. At 5 min after infection, aggregation of aerolysin-labeled GPI-anchored proteins showing tail-like formation was colocalized with swimming bacteria on the macrophage surface (Fig. 1). In contrast, no aggregation of CD48, which is a GPI-anchored protein, was observed at the same time point (Fig. 1). Similar results were obtained for other GPI-anchored proteins, such as CD55 (unpublished data). However, when one GPI-anchored protein, PrPC, was tested, colocalization of aggregated PrPC tail and swimming bacteria was observed (Fig. 2 A). Sometimes, several PrPC tails were observed from a single bacterium (Fig. 2 A). PrPC was also incorporated into macropinosomes containing wild-type strain, but not virB4 mutant, after 15 min incubation (Fig. 2 A).


Cellular prion protein promotes Brucella infection into macrophages.

Watarai M, Kim S, Erdenebaatar J, Makino S, Horiuchi M, Shirahata T, Sakaguchi S, Katamine S - J. Exp. Med. (2003)

Tail formation of GPI-anchored proteins on the site of swimming internalization. Bone marrow–derived macrophages were incubated with B. abortus for 5 or 15 min, and GPI-anchored proteins were localized by immunofluorescence as described in Materials and Methods. Merged images of the GFP (green) and TRITC (red) channels up and down with phase contrast images of the same field are shown. Cells were probed with aerolysin for GPI-anchored proteins and with anti-CD48. White arrows point to bacteria and blue arrow points to tail-like aggregation of GPI-anchored proteins.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196088&req=5

fig1: Tail formation of GPI-anchored proteins on the site of swimming internalization. Bone marrow–derived macrophages were incubated with B. abortus for 5 or 15 min, and GPI-anchored proteins were localized by immunofluorescence as described in Materials and Methods. Merged images of the GFP (green) and TRITC (red) channels up and down with phase contrast images of the same field are shown. Cells were probed with aerolysin for GPI-anchored proteins and with anti-CD48. White arrows point to bacteria and blue arrow points to tail-like aggregation of GPI-anchored proteins.
Mentions: Our previous results showed that GPI-anchored proteins were selectively incorporated into macropinosomes containing B. abortus (Fig. 1; reference 6). To investigate further the membrane sorting process, the distribution of GPI-anchored proteins during swimming internalization of B. abortus was analyzed. Aerolysin from Aeromonas hydrophila, which binds to the GPI moiety of GPI-anchored proteins on the cell surface (26), was used as probe for the detection of GPI-anchored proteins. At 5 min after infection, aggregation of aerolysin-labeled GPI-anchored proteins showing tail-like formation was colocalized with swimming bacteria on the macrophage surface (Fig. 1). In contrast, no aggregation of CD48, which is a GPI-anchored protein, was observed at the same time point (Fig. 1). Similar results were obtained for other GPI-anchored proteins, such as CD55 (unpublished data). However, when one GPI-anchored protein, PrPC, was tested, colocalization of aggregated PrPC tail and swimming bacteria was observed (Fig. 2 A). Sometimes, several PrPC tails were observed from a single bacterium (Fig. 2 A). PrPC was also incorporated into macropinosomes containing wild-type strain, but not virB4 mutant, after 15 min incubation (Fig. 2 A).

Bottom Line: Hsp60 reacted strongly with serum from human brucellosis patients and was exposed on the bacterial surface via a VirB complex-associated process.Under in vitro and in vivo conditions, Hsp60 of B. abortus bound to PrPC.These results indicate that signal transduction induced by the interaction between bacterial Hsp60 and PrPC on macrophages contributes to the establishment of B. abortus infection.

View Article: PubMed Central - PubMed

Affiliation: Department of Applied Veterinary Science, Obihiro University of Agriculture and Veterinary Medicine, Inada-cho, Obihiro-shi, Hokkaido 080-8555, Japan. watarai@obihiro.ac.jp

ABSTRACT
The products of the Brucella abortus virB gene locus, which are highly similar to conjugative DNA transfer system, enable the bacterium to replicate within macrophage vacuoles. The replicative phagosome is thought to be established by the interaction of a substrate of the VirB complex with macrophages, although the substrate and its host cellular target have not yet been identified. We report here that Hsp60, a member of the GroEL family of chaperonins, of B. abortus is capable of interacting directly or indirectly with cellular prion protein (PrPC) on host cells. Aggregation of PrPC tail-like formation was observed during bacterial swimming internalization into macrophages and PrPC was selectively incorporated into macropinosomes containing B. abortus. Hsp60 reacted strongly with serum from human brucellosis patients and was exposed on the bacterial surface via a VirB complex-associated process. Under in vitro and in vivo conditions, Hsp60 of B. abortus bound to PrPC. Hsp60 of B. abortus, expressed on the surface of Lactococcus lactis, promoted the aggregation of PrPC but not PrPC tail formation on macrophages. The PrPC deficiency prevented swimming internalization and intracellular replication of B. abortus, with the result that phagosomes bearing the bacteria were targeted into the endocytic network. These results indicate that signal transduction induced by the interaction between bacterial Hsp60 and PrPC on macrophages contributes to the establishment of B. abortus infection.

Show MeSH
Related in: MedlinePlus