RIAM Interacts with the Hematopoietic-Specific Adaptor Protein Gads and Forms a LAT-Independent Node of Signal Integration That Regulates Activation of PLC-γ1
Notice bibliographique
Résumé
Abstract TCR stimulation triggers the activation of protein tyrosine kinases resulting in phosphorylation of the adaptor protein LAT. SLP-76, interacts constitutively with PLC-γ1 and with the SH3 domain of Gads, which via its SH2 domain mediates inducible recruitment of SLP-76 and PLC-γ1 to LAT, upon T cell activation. PLC-γ1 hydrolyzes phosphatidylinositol-4, 5 bisphosphate [PI(4,5)P2], generating inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG), second messengers responsible for mediating intracellular calcium release and activation of downstream signals. The adaptor protein RIAM constitutively interacts with PLC-γ1 and is required for PLC-γ1 activation. RIAM is a multidomain protein with a small N-terminus proline-rich region, two coiled-coiled regions, sequential Ras association (RA) and pleckstrin homology (PH) domains, and a large C-terminus proline-rich region, which interacts with PLC-γ1. The RA domain of RIAM has specificity for Rap1-GTP whereas the PH domain binds to the PLC-γ1 substrate PI(4,5)P2. The RA-PH domain region of RIAM functions as a single structural unit and mediates translocation of RIAM to the plasma membrane upon T cell activation. Previously, we determined that RIAM deficiency results in impaired activation of PLC-γ1 in spite of the formation of the PLC-γ1-SLP-76-LAT complex, suggesting perhaps somewhat paradoxically, that PLC-γ1-SLP-76-LAT signalosome is not sufficient to mediate distal signaling in the absence of RIAM. This observation indicated that RIAM mediates its effects at a level distal to SLP-76-LAT or through a signaling pathway parallel but distinct from SLP-76-Gads-LAT. Here we investigated whether RIAM forms a signalosome parallel to PLC-γ1-SLP-76-Gads and whether such pathway might be involved in the activation of PLC-γ1. Using primary T lymphocytes and Jurkat T cells stimulated via TCR/CD3 and CD28 we determined that RIAM constitutively interacted with Gads as determined by immunoprecipitation with RIAM-specific antibody followed by Gads immunoblot. To determine whether the interaction between RIAM and Gads was direct, we employed an in vitro protein association assay. Glutathione S-transferase (GST) and GST-fusion protein of Gads were coupled to glutathione-sepharose and incubated with [35S]methionine-labeled RIAM or luciferase, as negative control. Gads bound to [35S]methionine-labeled RIAM indicating that RIAM interacts directly with Gads. We further examined domain-specific interaction of RIAM with endogenous Gads using GST fusion proteins of RIAM. We determined a constitutive interaction between Gads and GST fusion proteins of full-length RIAM or C-terminus region of RIAM. Although a number of tyrosine phosphorylated proteins were associated with the RIAM-Gads complex upon T cell activation, LAT was not detected among the components of this complex as determined by immunoblot with anti-phosphotyrosine-specific or LAT-specific antibodies. Using a GST fusion protein of the RA-PH domain of RIAM we determined that, surprisingly, Gads displayed activation-dependent interaction with the RA-PH domain, which mediates the recruitment of RIAM to the plasma membrane upon T cell activation. Furthermore, in addition to Gads, SLP-76 and PLC-γ1 were recruited to the RA-PH domain of RIAM in activated T cells. To determine whether RIAM and Gads had a synergistic effect on IL-2 transcription, we performed luciferase-based reporter assays using a reporter construct driven by the entire IL-2 promoter or by NFAT binding sequences. We found that RIAM and Gads had a synergistic effect on IL-2 and on NFAT-mediated transcriptional activation, which depends on PLC-γ1. Thus, via its C-terminus region, RIAM directly and constitutively interacts with Gads. In addition, via its RA-PH domain, RIAM mediates an activation-dependent interaction with Gads and serves as a docking site recruiting the PLC-γ1-SLP-76-Gads complex to the plasma membrane in a LAT-independent manner. These findings indicate a crosstalk between RIAM and SLP-76 in the activation of PLC-γ1 and reveal a previously unidentified, alternative signaling pathway leading to Gads-SLP-76 recruitment to the plasma membrane of activated T cells in a LAT-independent manner. Disclosures No relevant conflicts of interest to declare.
Récupéré en direct depuis OpenAlex et désinversé. Les résumés ne sont pas conservés dans cette base de données : les index inversés représentent 8,6 Go des 9,3 Go de texte de la base, et le serveur dispose de 13 Go libres.
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 ».