A mouse renin promoter containing the conserved decanucleotide element binds the same B-cell factors as an authentic immunoglobulin heavy chain promoter

AbstractA mouse renin-1 gene promoter fragment, normally inactive in B-cells, becomes a potent promoter in these cells after insertion of the highly conserved decanucleotide (dc/cd sequence) of immunoglobulin heavy and light chain promoters [(1987) EMBO J. 6, 1685–1690]. We observe retarded complexes of the same electrophoretic mobility when the cd-containing renin promoter fragment or an authentic immunoglobulin heavy chain promoter fragment is incubated with a nuclear extract from myeloma cells, suggesting that the renin promoter is activated due to its acquired ability to bind a B-cell-specific positive factor. No retarded complexes are observed with the original renin promoter fragment thus questioning the presence of a repressor as an explanation for its lack of activity in B-cells

Conferring a template-dependent polymerase activity to terminal deoxynucleotidyltransferase by mutations in the Loop1 region

Terminal deoxynucleotidyltransferase (Tdt) and DNA polymerase μ (pol μ) are two eukaryotic highly similar proteins involved in DNA processing and repair. Despite their high sequence identity, they differ widely in their activity: pol μ has a templated polymerase activity, whereas Tdt has a non-templated one. Loop1, first described when the Tdt structure was solved, has been invoked as the major structural determinant of this difference. Here we describe attempts to transform Tdt into pol μ with the minimal number of mutations in and around Loop1. First we describe the effect of mutations on six different positions chosen to destabilize Tdt Loop1 structure, either by alanine substitution or by deletion; they result at most in a reduction of Tdt activity, but adding Co++ restores most of this Tdt activity. However, a deletion of the entire Loop1 as in pol λ does confer a limited template-dependent polymerase behavior to Tdt while a chimera bearing an extended pol μ Loop1 reproduces pol μ behavior. Finally, 16 additional substitutions are reported, targeted at the two so-called ‘sequence determinant’ regions located just after Loop1 or underneath. Among them, the single-point mutant F401A displays a sequence-specific replicative polymerase phenotype that is stable upon Co++ addition. These results are discussed in light of the available crystal structures

Host Alternation Is Necessary to Maintain the Genome Stability of Rift Valley Fever Virus

Arthropod-borne viruses are transmitted among vertebrate hosts by insect vectors. Unusually, Rift Valley fever virus (RVFV) can also be transmitted by direct contacts of animals/humans with infectious tissues. What are the molecular mechanisms and evolutionary events leading to adopt one mode of transmission rather than the other? Viral replication is implied to be different in a vertebrate host and an invertebrate host. The alternating host cycle tends to limit virus evolution by adopting a compromise fitness level for replication in both hosts. To test this hypothesis, we used a cell culture model system to study the evolution of RVFV. We found that freeing RVFV from alternating replication in mammalian and mosquito cells led to large deletions in the NSs gene carrying the virulence factor. Resulting NSs-truncated viruses were able to protect mice from a challenge with a virulent RVFV. Thus, in nature, virulence is likely maintained by continuous alternating passages between vertebrates and insects. Thereby, depending on the mode of transmission adopted, the evolution of RVFV will be of major importance to predict the outcome of outbreaks

Etude de l'évolution et de l'expression du gene VcsA1 codant pour une prohormone de la glande sous-maxillaire de rat

PARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Contribution à l'analyse des mécanismes d'hypermutation des régions variables des immunoglobulines

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Incorporation de dUTP et hypermutation somatique des gènes des immunoglobulines

L hypermutation somatique (SHM) des gènes des immunoglobulines est le résultat d un processus initié par l enzyme AID (activation-induced cytidine deaminase), une cytidine-désaminase. Deux mécanismes différents sont responsables de l apparition des mutations au niveau des paires CG (phase I) et AT (phase II). Le mismatch U:G résultant de l action de l AID peut être répliqué, ce qui donnera des mutations de transition au niveau des paires CG, ou l uracile excisé, formant un site abasique qui, répliqué, résultera en des mutations de transition/transversion au niveau des paires CG. Alternativement, le mismatch peut être reconnu par le système de MMR (mismatch-repair), qui excise le brin contenant l uracile. C est pendant sa resynthèse par une ADN-polymérase que sont introduites les mutations au niveau des paires AT. Deux mécanismes ont été proposés pour rendre compte de cette sélectivité : des ADN-polymérases error-prone spécifiques, et/ou l incorporation de dUTP. Cependant, aucune donnée expérimentale n avait encore été obtenue quant à la possible contribution de ce dernier. Nous avons mis au point une méthode basée sur la LA-PCR (ligation-anchored PCR), nous permettant d identifier la présence d uracile dans les gènes VkOx de souris immunisées, à la fois dans le contexte de mismatchs U:G (résultants de l action de l AID), et de paires U:A (résultants de l incorporation de dUTP). Ces résultats montrent que le mécanisme d incorporation de dUTP est impliqué dans l hypermutation somatique, et pourrait constituer le mécanisme principal responsable de l apparition des mutations de phase IIPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF