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Ubiquitination and degradation of the Crohn’s Disease associated protein Nod2involves the E2 enzyme UBE2G2
Linköping University, Department of Clinical and Experimental Medicine, Surgery . Linköping University, Faculty of Health Sciences.
Institute for Medical Microbiology, Immunology and Hygiene, University of Cologne, Germany.
Östergötlands Läns Landsting, Centre of Surgery and Oncology, Department of Surgery in Östergötland. (Landstinget i Östergötland)
University of Toronto, Toronto, Canada.
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Background: Mutations predisposing to Crohn’s disease (CD) have been mapped to the CARD15/Nod2 locus,which encodes a cytoplasmic receptor hereafter referred to as Nod2, a member of the NACHT-LRR (NLR) familyof pattern recognition receptors. The binding of bacterial muramyl dipeptide (MDP) to Nod2 triggers theactivation of the nuclear factor-κB (NFκB) pathway, thereby inducing a number of pro-inflammatory genes. Themost common variant of Nod2 associated with CD is the frame shift mutation L1007fs, which results in atruncated form of the protein unable to respond to MDP. Despite active research, little is known about howNod2 is regulated. The aim of this study was to investigate if the cellular Nod2 protein level is regulated by theubiquitin-proteasome pathway.

Material and Methods/Results: Nod2 was shown to be subjected to rapid turnover in the colorectal cancer cellline SW480 as measured by immunoprecipitation following inhibition of protein synthesis with cyklohexamide.Immunoprecipitation of Nod2 also revealed co-precipitation of ubiquitin, suggesting that Nod2 wasubiquitinated. In line with this observation, inhibition of the proteasome using MG-132 or lactacystin, resultedin increased levels of Nod2 in the cells. UBE2G2, an E2 enzyme, thus conferring specificity of ubiquitin binding,was identified to have affinity for the CARD domain of Nod2. Activation of Nod2 with MDP enhanced itsubiquitination, and increasing amounts of UBE2G2 in the cells abrogated NFB-activation, suggesting thatubiquitination of Nod2 may be important for the resolution of the inflammatory signal.

Conclusion: Taken together, our results show that the cellular level of Nod2 protein is regulated via theubiquitin-proteasome pathway, suggesting that Nod2-driven inflammation may be resolved through rapiddegradation of Nod2. Consequently, not only polymorphisms in Nod2 directly, but also in genes regulatingNod2 protein levels may contribute to the susceptibility to Crohn’s disease.

National Category
Medical and Health Sciences
URN: urn:nbn:se:liu:diva-21099OAI: diva2:240584
Available from: 2009-09-29 Created: 2009-09-29 Last updated: 2010-01-14Bibliographically approved
In thesis
1. Mechanisms of bacterial-epithelial interaction in Crohn’s disease
Open this publication in new window or tab >>Mechanisms of bacterial-epithelial interaction in Crohn’s disease
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Crohn’s disease (CD) is believed to be initiated when an individual, who has agenetic predisposition either leading to a disturbance in the barrier functionand/or the innate immune system is exposed to triggering environmentalfactors, the most important being intraluminal bacteria. Genetic and functionalstudies have confirmed the Pattern-recognition receptors (PRRs), Nod2, TLR4and NALP3, as important mediators of the inflammatory process associatedwith disease progression. However, the mechanisms that link enteric bacteriaand barrier function in a background of genetic predisposition to CD are justbeginning to emerge. The general aim of this thesis was therefore to morethoroughly investigate the mechanisms of bacterial-epithelial interaction in CD.

Here we present evidence suggesting that the small bowel is able to inducetranscytosis of antigens after short term exposure to Yersinia pseudotuberculosis. This suggests that small bowel enterocytes are able toattain follicle associated epithelial (FAE) abilities and contribute to the barrierdysfunction observed in CD. Furthermore we report a positive effect of anti-TNFα treatment (infliximab) on the translocation of adherent invasive E.coli (AIEC) across the colonic mucosa of patients suffering from severe CD.

We also confirm the importance of the Nod-like receptors (NLRs) in thepathogenesis of CD by showing that combined polymorphisms in the genesencoding NALP3 and CARD8 confer susceptibility to CD among Swedish menand in addition to previous published results add a gender aspect on thegenotype-phenotype relationship in CD. Finally, we show that Nod2 is rapidlysubjected to ubiquitination followed by proteasomal degradation, henceproviding important clues about how NLR regulation might occur in the cell,suggesting that the ubiquitin-proteasome pathway is an important factor toconsider in the development of the disease.

In conclusion we report novel insights into the bacterial-epithelial interactionsoccurring in CD and contribute important clues about the origin of this disease.ISBN: 978-

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2009. 71 p.
Linköping University Medical Dissertations, ISSN 0345-0082 ; 1113
National Category
Medical and Health Sciences
urn:nbn:se:liu:diva-21104 (URN)978-91-7393-667-5 (ISBN)
Public defence
2009-04-17, Berzeliussalen, Hälsouniversitetet, Campus US, Linköpings Universitet, Linköping, 13:00 (English)
Available from: 2009-09-29 Created: 2009-09-29 Last updated: 2009-09-29Bibliographically approved

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