A 3' → 5' XPB helicase defect in repair/transcription factor TFIIH of xeroderma pigmentosum group B affects both DNA repair and transcription

XPB is a subunit of the basal transcription factor TFIIH, which is also involved in nucleotide excision repair (NER) and potentially in cell cycle regulation. A frameshift mutation in the 3'-end of the XPB gene is responsible for a concurrence of two disorders: xeroderma pigmentosum (XP) and Cockayne's syndrome (CS). We have isolated TFIIH from cells derived from a patient (XP11BE) who carries this frameshift mutation (TFI-IHmut) and from the mother of this patient (TFIIHwt) to determine the biochemical consequences of the mutation. Although identical in composition and stoichiometry to TFIIHwt, TFIIHmut shows a reduced 3' → 5' XPB helicase activity. A decrease in helicase and DNA-dependent ATPase activities was also observed with the mutated recombinant XPB protein. The XPB mutation causes a severe NER defect. In addition, we provide evidence for a decrease in basal transcription activity in vitro. The latter defect may provide an explanation for many of the XP and CS symptoms that are difficult to rationalize based solely on an NER defect. Thus, this work presents the first detailed analysis of a naturally occurring mutation in a basal transcription factor and supports the concept that the combined XP/CS clinical entity is actually the result of a combined transcription/repair deficiency.</p

A 3' → 5' XPB helicase defect in repair/transcription factor TFIIH of xeroderma pigmentosum group B affects both DNA repair and transcription

XPB is a subunit of the basal transcription factor TFIIH, which is also involved in nucleotide excision repair (NER) and potentially in cell cycle regulation. A frameshift mutation in the 3'-end of the XPB gene is responsible for a concurrence of two disorders: xeroderma pigmentosum (XP) and Cockayne's syndrome (CS). We have isolated TFIIH from cells derived from a patient (XP11BE) who carries this frameshift mutation (TFI-IHmut) and from the mother of this patient (TFIIHwt) to determine the biochemical consequences of the mutation. Although identical in composition and stoichiometry to TFIIHwt, TFIIHmut shows a reduced 3' → 5' XPB helicase activity. A decrease in helicase and DNA-dependent ATPase activities was also observed with the mutated recombinant XPB protein. The XPB mutation causes a severe NER defect. In addition, we provide evidence for a decrease in basal transcription activity in vitro. The latter defect may provide an explanation for many of the XP and CS symptoms that are difficult to rationalize based solely on an NER defect. Thus, this work presents the first detailed analysis of a naturally occurring mutation in a basal transcription factor and supports the concept that the combined XP/CS clinical entity is actually the result of a combined transcription/repair deficiency.</p

Muscle inactivation of mTOR causes metabolic and dystrophin defects leading to severe myopathy

Mammalian target of rapamycin (mTOR) is a key regulator of cell growth that associates with raptor and rictor to form the mTOR complex 1 (mTORC1) and mTORC2, respectively. Raptor is required for oxidative muscle integrity, whereas rictor is dispensable. In this study, we show that muscle-specific inactivation of mTOR leads to severe myopathy, resulting in premature death. mTOR-deficient muscles display metabolic changes similar to those observed in muscles lacking raptor, including impaired oxidative metabolism, altered mitochondrial regulation, and glycogen accumulation associated with protein kinase B/Akt hyperactivation. In addition, mTOR-deficient muscles exhibit increased basal glucose uptake, whereas whole body glucose homeostasis is essentially maintained. Importantly, loss of mTOR exacerbates the myopathic features in both slow oxidative and fast glycolytic muscles. Moreover, mTOR but not raptor and rictor deficiency leads to reduced muscle dystrophin content. We provide evidence that mTOR controls dystrophin transcription in a cell-autonomous, rapamycin-resistant, and kinase-independent manner. Collectively, our results demonstrate that mTOR acts mainly via mTORC1, whereas regulation of dystrophin is raptor and rictor independent

Muscle inactivation of mTOR causes metabolic and dystrophin defects leading to severe myopathy

mTor, acting mainly via mTORC1, controls dystrophin transcription in a raptor- and rictor-independent mechanism

Etude d'un facteur stimulant la transcription en se liant aux sequences amont du promoteur tardif majeur de l'Adenovirus-2

SIGLECNRS T Bordereau / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Expression inEscherichia coli: Purification and properties of the recombinant human general transcription factor rTFII B

International audienc

Etude du rôle des modifications post-traductionnelles des histones dans l’acclimatation embryonnaire à la chaleur chez le poulet

L’environnement précoce des individus est capable d’impacter l’expression des gènes à long terme via des altérations de leur épigénome. Par exemple chez Drosophila melanogaster l’exposition à un stress thermique au cours de l’embryogenèse induit des modifications épigénétiques visibles à l’âge adulte via le complexe PRC2 (Polycomb Repressive Complex 2) (pour revue Steffen and Ringrose 2014). Une marque d’histones modulée par PRC2 est la tri-méthylation de la lysine 27 sur l’histone H3 (H3K27Me3). Cette marque est décrite comme ayant un rôle dans la mémoire d’un état répressif sous l’influence de l’environnement. Notamment chez Arabidopsis thaliana, suite à l’exposition au froid de l’hiver cette marque maintient la répression d’un répresseur floral pour permettre la floraison (Coustham et al. 2012). Dans un contexte de réchauffement climatique, notre équipe a étudié les effets d’un protocole expérimental permettant d’acclimater les poulets à la chaleur pendant l’embryogénèse (Piestun et al., 2008 ; Loyau et al., 2013). En effet, les animaux sélectionnés pour leurs performances de croissance sont peu résistants aux variations de températures. Le traitement d’acclimatation embryonnaire à la chaleur (TAEC) correspond à une augmentation cyclique de la température et de l’humidité relative (HR) pendant l’incubation des jours E7 à E16 de l’embryogénèse (de 37,8°C et 55% HR à 39,5°C et 65% HR pendant 12h). Cette modification de l’environnement précoce des individus altère peu l’éclosabilité et améliore le taux de survie lors d’un coup de chaleur survenant à l’âge d’abattage (J35). Cette thermotolérance accrue est associée de changements physiologiques, métaboliques et d’expressions de gènes à 5 semaines (Piestun et al., 2011, et Loyau et al., 2013, 2014). Nos recherches reposent sur l’hypothèse que les différences d’expressions de gènes observées chez les individus acclimatés peuvent être induites par des modifications épigénétiques survenant durant le TAEC et qui perdurent au cours du développement. Les travaux présentés porteront sur la mise au point de l’immunoprécipitation de la chromatine (ChIP) sur deux tissus de poulet afin de cartographier H3K27Me3

Variations épigénétiques en réponse à la chaleur chez le poulet de chair

National audienc

Genomic structure of the human TATA-box-binding protein (TBP)

International audienc

DNA repair helicase:A component of BTF2 (TFIIH) basic transcription factor

The human BTF2 basic transcription factor (also called TFIIH), which is similar to the δ factor in rat and factor b in yeast, is required for class II gene transcription. A strand displacement assay was used to show that highly purified preparation of BTF2 had an adenosine triphosphate-dependent DNA helicase activity, in addition to the previously characterized carboxyl-terminal domain kinase activity. Amino acid sequence analysis of the tryptic digest generated from the 89-kilodalton subunit of BTF2 indicated that this polypeptide corresponded to the ERCC-3 gene product, a presumed helicase implicated in the human DNA excision repair disorders xeroderma pigmentosum and Cockayne's syndrome. These findings suggest that transcription and nucleotide excision repair may share common factors and hence may be considered to be functionally related.</p