Cellular targets of Pseudomonas aeruginosa toxin Exoenzyme S [Elektronisk resurs]
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Henriksson, Maria, 1971- (författare)
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Hallberg, Bengt (preses)
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Rönnstrand, Lars (opponent)
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Umeå universitet Medicinska fakulteten (utgivare)
- Publicerad: Umeå universitet, 2003
- Engelska 51
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Serie: Umeå University medical dissertations, 0346-6612 0346-6612
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- Relaterad länk:
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http://www.umu.se/ (Värdpublikation)
Sammanfattning
Ämnesord
Stäng
- Pseudomonas aeruginosa is an opportunistic pathogen that can cause life-threatening infections in immunocompromised patients. It uses a type III secretion dependent mechanism to translocate toxic effector proteins directly into the eukaryotic cell. The enzymatic activity of two of these toxins, Exoenzyme S (ExoS) and Exoenzyme T (ExoT), have been studied in this thesis. ExoS is a bi-functional toxin known to contain a C-terminal ADP-ribosyltransferase activity, which has been shown to modify members of the Ras family in vitro. The N-terminal of ExoS contains a GTPase Activating Protein (GAP) domain, which shows specificity towards Rho proteins in vitro. ExoT shows high homology (76%) towards ExoS and has also been reported to contain ADP-ribosyltransferase activity in vitro . To study the biological effect of the two toxins, we inserted ExoS or ExoT into eukaryotic cells using the heterologous type III secretion system of Yersinia pseudotuberculosis . We found that Ras was ADP-ribosylated in vivo and this modification altered the ratio of GTP/GDP bound directly to Ras. We also found that ExoS could ADP-ribosylate several members of the Ras superfamily in vivo , modulating the activity of those proteins. In contrast, ExoT showed no ADP-ribosylation activity towards any of the GTPases tested. This suggests that ExoS is the major ADP-ribosyltransferase modulating small GTPase function encoded by P. aeruginosa . Furthermore, we have demonstrated that the GAP activity of ExoS abolishes the activation of RhoA, Cdc42 and Rap1 in vivo , and that ExoT shows GAP activity towards RhoA in vitro . The ADP-ribosyltransferase activity of ExoS is dependent on the eukaryotic protein 14-3-3. 14-3-3 proteins interact with ExoS in a phospho-independent manner. We identified the amino acids 424 DALDL 428 on ExoS to be necessary for the specific interaction between ExoS and 14-3-3. Deletion of these five amino acids abolishes the ADP-ribosylation of Ras and hence the cytotoxic effect of P. aeruginosa on cells. Thus the 14-3-3 binding motif on ExoS appears to be critical for both the ADP-ribosylation activity and the cytotoxic action of ExoS in vivo .
Ämnesord
- Medical and Health Sciences (hsv)
- Basic Medicine (hsv)
- Cell and Molecular Biology (hsv)
- Medicin och hälsovetenskap (hsv)
- Medicinska och farmaceutiska grundvetenskaper (hsv)
- Cell- och molekylärbiologi (hsv)
- MEDICINE (svep)
- Morphology, cell biology, pathology (svep)
- Cell biology (svep)
- MEDICIN (svep)
- Morfologi, cellbiologi, patologi (svep)
- Cellbiologi (svep)
- molekylär cellbiologi (umu)
- Molecular Cellbiology (umu)
Genre
- government publication (marcgt)
Indexterm och SAB-rubrik
- Cell biology
- Pseudomonas aeruginosa
- ADP-ribosylation
- GAP
- Ras superfamily
- NAD
- ExoS
- 14-3-3
- Cellbiologi
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