| dc.description | Legionella pneumophila is the facultative intracellular bacterium that causes an acute pneumonia known as Legionnaires' disease. During the infection of host cells, one of the most abundant proteins expressed by L. pneumophila is HtpB, a member of the 60-kDa chaperonin family of molecular chaperones. L. pneumophila uses HtpB to attach to and invade HeLa cells, but the continued synthesis and release of HtpB by internalized bacteria suggests that HtpB may also play a role in the intracellular events directed by L. pneumophila. Intracellular events leading to the generation of the L. pneumophila replicative vacuole include recruitment of mitochondria and endoplasmic reticulum (ER)-derived vesicles, evasion of fusion with lysosomes, and association with the ER. To investigate whether HtpB contributes to these events, the intracellular trafficking of HtpB-coated beads in Chinese hamster ovary (CHO) and U937 cells was examined by fluorescence and electron microscopy. Internalized HtpB-coated beads mimicked the ability of virulent L. pneumophila to recruit mitochondria and transiently induce actin rearrangements, but did not mimic the ability of L. pneumophila to avoid fusion with lysosomes. cAMP translocation assay and immunogold electron microscopy were performed to investigate whether HtpB has access to the host cytoplasm after being released into the L. pneumophila-containing vacuole (LCV). By these methods, HtpB was found to be delivered into the host cytoplasm where it associates with the cytoplasmic face of the LCV. CHO cells expressing HtpB displayed altered actin organization, demonstrating a role for HtpB in this compartment. HtpB and GroEL co-sedimented with and induced depolymerization of F-actin in vitro, suggesting a direct interaction between bacterial chaperonins and actin. This interaction was excluded as the underlying mechanism for the HtpB-mediated cytoskeletal rearrangements since ectopically expressed GroEL was unable to alter actin organization in CHO cells. In summary, this study reports the translocation of HtpB into the host cytoplasm, and the ability of HtpB to attract mitochondria and remodel the actin cytoskeleton in mammalian cells. These novel abilities described for HtpB may contribute to the intracellular establishment of L. pneumophila, and provide support for the concept of molecular chaperones as virulence factors. | en_US |
