RUNX1-dependent RAG1 deposition instigates human TCR-δ locus rearrangement

V(D)J recombination of TCR loci is regulated by chromatin accessibility to RAG1/2 proteins, rendering RAG1/2 targeting a potentially important regulator of lymphoid differentiation. We show that within the human TCR-α/δ locus

Fonctionnement et régulation des systèmes respiratoires triméthylamine N-oxyde (TMAO) réductase chez Escherichia coli et Shewanella oneidensis

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Reconstitution of the Trimethylamine Oxide Reductase Regulatory Elements of Shewanella oneidensis in Escherichia coli

Several bacteria can grow by using small organic compounds such as trimethylamine oxide (TMAO) as electron acceptors. In Shewanella species, the TMAO reductase respiratory system is encoded by the torECAD operon. We showed that production of the TMAO reductase of S. oneidensis was induced by TMAO and repressed by oxygen, and we noticed that a three-gene cluster (torSTR) encoding a complex two-component regulatory system was present downstream of the torECAD operon. We introduced the torSTR gene cluster into Escherichia coli and showed that this regulatory gene cluster is involved in TMAO induction of the torE promoter but plays no role in the oxygen control. The TorR response regulator was purified, and gel shift and footprinting experiments revealed that TorR binds to a single region located about 70 bases upstream of the transcription start site of the tor structural operon. By deletion analysis, we confirmed that the TorR operator site is required for induction of the tor structural promoter. As the TMAO regulatory system of S. oneidensis is homologous to that of E. coli, we investigated a possible complementation between the TMAO regulatory components of the two bacteria. Interestingly, TorS(ec), the TMAO sensor of E. coli, was able to transphosphorylate TorR(so), the TMAO response regulator of S. oneidensis

Genes Regulated by TorR, the Trimethylamine Oxide Response Regulator of Shewanella oneidensis

The torECAD operon encoding the trimethylamine oxide (TMAO) respiratory system of Shewanella oneidensis is positively controlled by the TorS/TorR two-component system when TMAO is available. Activation of the tor operon occurs upon binding of the phosphorylated response regulator TorR to a single operator site containing the direct repeat nucleotide sequence TTCATAN(4)TTCATA. Here we show that the replacement of any nucleotide of one TTCATA hexamer prevented TorR binding in vitro, meaning that TorR specifically interacts with this DNA target. Identical direct repeat sequences were found in the promoter regions of torR and of the new gene torF (SO4694), and they allowed TorR binding to both promoters. Real-time PCR experiments revealed that torR is negatively autoregulated, whereas torF is strongly induced by TorR in response to TMAO. Transcription start site location and footprinting analysis indicate that the operator site at torR overlaps the promoter −10 box, whereas the operator site at torF is centered at −74 bp from the start site, in agreement with the opposite role of TorR in the regulation of the two genes. Since torF and torECAD are positively coregulated by TorR, we propose that the TorF protein plays a role related to TMAO respiration

Sequential role of RAD51 paralog complexes in replication fork remodeling and restart

Homologous recombination (HR) factors were recently implicated in DNA replication fork remodeling and protection. While maintaining genome stability, HR-mediated fork remodeling promotes cancer chemoresistance, by as-yet elusive mechanisms. Five HR cofactors - the RAD51 paralogs RAD51B, RAD51C, RAD51D, XRCC2 and XRCC3 - recently emerged as crucial tumor suppressors. Albeit extensively characterized in DNA repair, their role in replication has not been addressed systematically. Here, we identify all RAD51 paralogs while screening for modulators of RAD51 recombinase upon replication stress. Single-molecule analysis of fork progression and architecture in isogenic cellular systems shows that the BCDX2 subcomplex restrains fork progression upon stress, promoting fork reversal. Accordingly, BCDX2 primes unscheduled degradation of reversed forks in BRCA2-defective cells, boosting genomic instability. Conversely, the CX3 subcomplex is dispensable for fork reversal, but mediates efficient restart of reversed forks. We propose that RAD51 paralogs sequentially orchestrate clinically relevant transactions at replication forks, cooperatively promoting fork remodeling and restart

Fit αβ T-Cell Receptor suppresses leukemogenesis of Pten-deficient thymocytes.

International audienceSignaling through the αβT cell receptor (TCR) is a crucial determinant of T-cell fate and can induce two opposite outcomes during thymocyte development: cell death or survival and differentiation. To date, the role played by T cell receptor in the oncogenic transformation of developing T-cells remains unclear. Here we show that human primary T-cell acute lymphoblastic leukemias expressing a αβT cell receptor are frequently deficient for phosphatase and tensin homolog protein (PTEN), and fail to strongly respond to T cell receptor activation. Using Pten-deficient T-cell acute lymphoblastic leukemia mouse models, we confirm that T cell receptor signaling is involved in leukemogenesis. We show that abrogation of T cell receptor expression accelerated tumor onset, while enforced expression of a fit transgenic T cell receptor led to the development of T cell receptor-negative lymphoma and delayed tumorigenesis. We further demonstrate that pre-tumoral Pten-deficient thymocytes harboring fit T cell receptors undergo early clonal deletion, thus preventing their malignant transformation; while cells with unfit T cell receptors that should normally be deleted during positive selection, pass selection and develop T-cell acute lymphoblastic leukemias. Altogether, our data show that fit T cell receptor signaling suppresses tumor development mediated by Pten loss-of-function and pinpoint to a role of Pten in positive selection

Sequential role of RAD51 paralog complexes in replication fork remodeling and restart

International audienceHomologous recombination (HR) factors were recently implicated in DNA replication fork remodeling and protection. While maintaining genome stability, HR-mediated fork remodeling promotes cancer chemoresistance, by as-yet elusive mechanisms. Five HR cofactors - the RAD51 paralogs RAD51B, RAD51C, RAD51D, XRCC2 and XRCC3 - recently emerged as crucial tumor suppressors. Albeit extensively characterized in DNA repair, their role in replication has not been addressed systematically. Here, we identify all RAD51 paralogs while screening for modulators of RAD51 recombinase upon replication stress. Single-molecule analysis of fork progression and architecture in isogenic cellular systems shows that the BCDX2 subcomplex restrains fork progression upon stress, promoting fork reversal. Accordingly, BCDX2 primes unscheduled degradation of reversed forks in BRCA2-defective cells, boosting genomic instability. Conversely, the CX3 subcomplex is dispensable for fork reversal, but mediates efficient restart of reversed forks. We propose that RAD51 paralogs sequentially orchestrate clinically relevant transactions at replication forks, cooperatively promoting fork remodeling and restart