Histone Demethylases as Modulators of Hematopoietic Cell Fate.
Notice bibliographique
Résumé
Abstract Abstract 393 Epigenetic modifications influence cell fate decisions, such as stem cell self-renewal and differentiation, both in normal and leukemic stem cells (LSC). On this premise, an expression profile study of the emerging class of histone demethylating enzymes (HDE) in stem cell enriched populations was designed. Transcripts of all these enzymes (n=27) were detected in isolated HSC populations (frequency 1:2) from fetal liver and bone marrow. Similar results were obtained in HoxA9-Meis1 induced leukemia, with an inverse correlation between LSC content and the HDE Jarid1b expression level. As a functional follow-up, systematic knockdown of known or presumed HDE was undertaken in primary HSC using a RNA interference (RNAi) based approach. Five sh-RNA were designed and generated against every HDE and cloned into a retroviral vector, containing a GFP reporter gene. The screen was set as a loss-of-function assay, where sorted CD150+CD48-Lin- cells were co-cultured on retroviral producers for 5 days, in a 96-well setting, and a fraction of each well was then transplanted into 2 mice (day 0), along with competitor cells. The remaining cell fraction served to asses gene transfer by GFP epifluorescence measurements, and RNA isolated from sorted GFP+ cells was used to evaluate gene knockdown levels by Q-PCR analysis. Blood reconstitution was evaluated at regular intervals, tracking the contribution of the donor CD45.1+ transduced (GFP+) cells to recipient hematopoiesis over time. As baseline references, sh-RNA to luciferase (no effect) and the histone acetyl transferase myst3 (stem cell loss) were used, as well as hoxb4 over-expression (stem cell expansion). In the primary screen, RNAi against jarid1a, lsd1 and an arginine demethylating enzyme, resulted in a progressive decline in blood reconstitution levels from the transduced cells, ranging from a 50% to 80% reduction of the GFP content within the graft. On the other hand, knockdown of Jarid1b conferred an in vivo competitive advantage to the transduced cells. As a validation procedure, a 7 day culture step was introduced to the assay, hence increasing the signal to noise ratio, to better detect an increase in HSC activity, as recently reported. After a week in vitro (day 7), 1/8 equivalents of single well cultures were transplanted into 3 independent mice, and blood reconstitution levels serially assessed. Cells transduced with sh-RNA against jarid1b (3 constructs out of 6) contributed more significantly to host hematopoiesis than sh-RNA luciferase transduced cells (58±16% vs 26±3% GFP), or hoxb4 over-expressing cells (37±2% GFP), at comparable gene transfer rates, at the 14 week time point and beyond (3 independent experiments). In long-term recipients, differentiation potential of these cells was preserved, as evidenced by CD4+CD8+ thymic cells, B220+ splenic cells and CD11b+ bone marrow cells in the GFP positive contingent. Clonality studies on DNA isolated from these sorted populations are ongoing. There were no cases of leukemic transformation in all of the transplant recipients (n>30). In conclusion, this RNAi based study performed on primary hematopoietic cells links histone demethylases to modulations in cell fate. An opposite role seems to emerge among the H3K4me3/me2 HDE JARID1a and JARID1b regarding HSC activity, yet these proteins are highly homologous. Sequence alignments pinpoint to differences within the C-terminal portion, containing a PHD domain, and interestingly, dysregulation of this chromatin-binding PHD finger (NUP98-JARID1a) has been recently shown to play a direct role in leukemogenesis 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 machine sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Le volet Gemma est une étiquette directe du modèle pour chaque travail de la base, lue sur la notice réduite au titre. Le volet Codex est un classifieur appris des 10 348 étiquettes directes de Codex et calibré sur les taux pondérés de l'échantillon; les champs sans appui suffisant ne portent aucun appel Codex. Le mode candidate est l'union des deux volets; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont pas des étiquettes humaines.
Scores du classifieur distillé par catégorie (deux têtes)
| 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) | 0,001 | 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 source (Gemma direct ou Codex distillé), 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 ».