Elucidating the Role of SYK in the Intestinal Epithelium
Notice bibliographique
Résumé
Background: Inflammatory Bowel Disease (IBD) involves chronic and recurring inflammation of the gastrointestinal tract. We identified several patients presenting with immune dysregulation and systemic inflammation, including inflammation of the gut. Analysis of Whole Exome Sequencing data from pediatric IBD patients has identified five rare protein coding Spleen Tyrosine Kinase (SYK) variants. Functional studies have demonstrated increased kinase activity implicating them in disease pathophysiology. While the role of SYK is well characterized in immune cells, it is not understood what role it plays in the intestinal epithelium. Aims: The primary aim of this project is to understand the role SYK plays in the intestinal epithelium, what molecular pathways it signals through, and how its dysregulation leads to intestinal disease. Methods: To understand SYKs role in the intestinal epithelium, a BioID screen was used to identify novel SYK interacting proteins. BioID was done with wild type SYK as well as the patient derived variants in HEK293 cells. BioID hits were validated using co-immunoprecipitation (CoIP) and proximity ligation assay (PLA). Further functional analysis was done in Caco2 cells, an intestinal epithelial cell model, with western blot, ELISA, and luciferase assays to assess the effect of SYK activity on downstream inflammatory pathways. Finally, we investigated the effect of small molecule kinase inhibitors to explore possible novel treatment options. Results: The BioID screen revealed several high confidence interactors, with dysregulated interactions in the patient variants. USP25, a ubiquitin specific peptidase that is widely expressed, including in the intestinal epithelium, was identified as one of the strongest hits based on peptide counts. The interaction was validated with CoIP and PLA, and further analysis showed SYK activation decreased USP25 protein levels by up to eight-fold. The secretion of the proinflammatory chemokine, IL-8, in response to lipopolysaccharide (LPS) was found to be almost twice as high in Caco2 cells with patient SYK variants relative to WT. Cells expressing the patient variants of SYK showed increased NFkB and AP1 levels, indicating increased inflammatory gene expression when stimulated with 100ng/mL LPS. NFkB was also active for longer in cells expressing the patient variant of SYK post stimulation, with WT cells returning to baseline in under half an hour, while the patient variant stayed in an inflamed state for 36 to 48 hours after LPS was removed. Conclusions: Increased SYK activity is associated with the degradation of USP25. USP25 is known to cleave ubiquitin off TNF receptor associated factors (TRAFs). TRAFs are activated downstream of innate immune receptors such as TLR4, and need to be ubiquitinated for activation. USP25, which is responsible for the deubiquitination of these TRAFs, acts as a negative regulator of these pathways. The hyperactive SYK variants degrade USP25 and impair the cells’ ability to switch off inflammatory signaling pathways, indicated by the cells elongated period in an inflamed state. This research is funded by grants from Canadian Institute for Health Research (CIHR) and the Leona M. and Harry B. Helmsley Charitable Trust. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
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Comment cette classification a été obtenuedéplier
Prédiction distillée sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Apprise à partir de 10 348 étiquettes directes de Codex et de 10 348 étiquettes directes de Gemma. Le mode candidate est l'union des têtes enseignantes seuillées; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont ni des étiquettes humaines ni des étiquettes directes de modèles de pointe.
Scores Codex et Gemma par catégorie
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,000 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,000 |
| Bibliométrie | 0,000 | 0,000 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,000 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,000 | 0,000 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule tête enseignante, pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».