Abstract 1234: Erk-dependent phosphorylation of Notch intracellular domain (NIC) correlates with increased NIC-dependent transcription
Bibliographic record
Abstract
Abstract Background. Recent studies have suggested that the ability of Ras to transform cells depends on cooperation with the Notch signaling pathway. However, the molecular and cellular mechanisms involved in this cooperation remain unknown. Aim. To evaluate the impact of the Mek/Erk pathway, downstream of Ras, on Notch signaling. Methods. HEK293T and human pancreatic cancer cells MIA PaCa-2 were used. Mek/Erk activity was down-regulated by treatment with the specific inhibitor U0126 (10 µM). Transfections or retroviral infections of a constitutive active form of KRas or Mek (MekCA) were used to increase Mek/Erk activity. NIC-dependent transcription was measured by luciferase assays using a specific (CSL) luciferase reporter gene. Results. 1- Transient transfection of an activated KRas stimulated by 1.7-fold NIC-dependent transcription. Transfection of MekCA increased by 4.3-fold NIC-dependent transcription while a wild-type form of Mek (MekWT) was without effect. In parallel, treatment with U0126 reduced by 60% the basal NIC-dependent transcription and completely blocked the KRas- and MekCA-induced NIC-dependent transcription. This negative effect of U0126 on NIC-dependent transcription correlated with decreased nuclear staining of the NIC target gene Hes1. 2- Western blot analysis revealed higher molecular weight forms of NIC when cells were transfected/infected with MekCA as compared to control MekWT-transfected/infected cells. U0126 treatment prevented the MekCA-induced decreased in mobility shift of NIC suggesting that NIC is subjected to Mek-dependent post-translational modifications in vivo. 3- Immunoprecipitation followed by in vitro phosphatase assay revealed that these MekCA-induced higher molecular weight forms of NIC were phosphorylated forms of NIC. 4- By in vitro kinase assays, we demonstrated that NIC worked as Erk substrate. 5- Finally, following in vivo phospholabeling, we observed that NIC immunoprecipitated from MekCA-transfected cells was more phosphorylated as compared to NIC immunoprecipitated from MekWT-transfected cells. Conclusion. Our results provide the novel discovery that Mek/Erk signaling promotes NIC phosphorylation which correlates with increased NIC-dependent transcription. Our discovery thus provides a possible cellular mechanism by which Ras and Notch cooperate to promote cellular transformation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1234.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.007 | 0.002 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".