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Nuclear targeting of Porphyromonas gingivalis W50 protease in epithelial cells

Record type: Article

Porphyromonas gingivalis is an important pathogen associated with destructive periodontal disease and is able to invade the epithelial cell barrier. Its cysteine proteases are recognized as major virulence factors, and in this study, we examined the interaction of the arginine-specific protease with epithelial cells in culture. Three cell lines (KB, HeLa, and SCC4) were incubated with strain W50 culture supernatant; stained with monoclonal antibody 1A1, which recognizes an epitope on the adhesin (ß) component of the cysteine protease-adhesin ({alpha}/ß) heterodimer; and viewed using immunofluorescence microscopy. Within 1 h, the protease traversed the plasma membrane and was localized around the nucleus before becoming concentrated in the cytoplasm after 24 to 48 h. In contrast, the purified arginine-specific heterodimeric protease (HRgpA) rapidly entered the nucleus within 15 to 30 min. This nuclear targeting (i) was seen with active and N{alpha}-p-tosyl-L-lysine chloromethyl ketone (TLCK)-inactivated HRgpA, indicating it was independent of the proteolytic activity; (ii) occurred at both 4 and 37°C; and (iii) failed to occur with the monomeric protease (RgpAcat), indicating the importance of the adhesin chain of the HRgpA protease to this process. Rapid cell entry was also observed with recombinant catalytic ({alpha}) and adhesin (ß) chains, with the latter again targeting the nuclear area. After 48 h of incubation with HRgpA, significant dose-dependent stimulation of metabolic activity was observed (measured by reduction of 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide), and a doubling of mitotic activity combined with the presence of apoptotic cells indicated that HRgpA may interfere with cell cycle control mechanisms. These effects were seen with both active and TLCK-inactivated protease, confirming that they were not dependent on proteolytic activity, and thus provide new insights into the functioning of this P. gingivalis protease.

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Citation

Scragg, M.A., Alsam, A., Rangarajan, M., Slaney, J.M., Shepherd, P., Williams, D.M. and Curtis, M.A. (2002) Nuclear targeting of Porphyromonas gingivalis W50 protease in epithelial cells Infection and Immunity, 70, (10), pp. 5740-5750. (doi:10.1128/IAI.70.10.5740-5750.2002).

More information

Published date: October 2002

Identifiers

Local EPrints ID: 48430
URI: http://eprints.soton.ac.uk/id/eprint/48430
ISSN: 0019-9567
PURE UUID: 85c5bc14-1bf8-438f-84ca-0659a0a0546d

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Date deposited: 21 Sep 2007
Last modified: 17 Jul 2017 14:59

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Contributors

Author: M.A. Scragg
Author: A. Alsam
Author: M. Rangarajan
Author: J.M. Slaney
Author: P. Shepherd
Author: D.M. Williams
Author: M.A. Curtis

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