Synthesis and triplex binding properties of oligonucleotides containing a novel nucleobase

The thiazolo-indole compound 1 bearing the complementary donor-acceptor-donor sites (dad) was designed for specific recognition of an AT inverted base pair in pyrimidine triple helix motif. It was successfully incorporated into 14-mer oligonucleotide using a serinol unit as sugar derivative. The triple helix hybridization studies were examined by means of thermal denaturation experiments with a 26-mer DNA duplex containing the AT inverted base pair.     

Origin and history of the IVS-I-110 and codon 39 beta-thalassemia mutations in the Lebanese population

Using restriction fragment length polymorphisms (RFLPs) and sequence haplotype analysis, we studied the chromosomal background of the beta-globin gene in 31 unrelated Lebanese IVS-I-110 or codon 39 (Cd39) subjects, and five normal betaAbeta/A individuals. Our results are compared with those from similar studies in other parts of the Mediterranean in an attempt to provide insights into historical patterns of selection and disease. The great majority of the Lebanese chromosomes with the IVS-I-110 mutation are associated with the RFLP haplotype I and sequence haplotype HT1, which is probably the ancestral structure on which the mutation first emerged. The remainder of the IVS-I-110 alleles are linked to the 5'-subhaplotype 12 RFLP haplotype and/or HTR sequence haplotype. In contrast, in Turkey, IVS-I-110 is associated with six distinct sequence haplotypes and four distinct RFLP haplotypes, suggesting that the mutation probably emerged there. The diversity of sequence haplotypes described in Turkey was probably generated through recombination or gene conversion events with the most frequent betaA autochthonous structures. Our data on Lebanese betaA chromosomes and Algerian betaA chromosomes, along with previously described Turkish betaA chromosomes, strengthen this hypothesis. Following its emergence in Turkey, the IVS-1-110 mutation was probably introduced to Lebanon later, by migration or settlements. Cd39 demonstrates a remarkable level of sequence and RFLP haplotype heterogeneity in Algeria, in contrast to its relative homogeneity in Turkish samples. However, its rarity in the Near East, and more specifically in Lebanon, does not allow us to draw any conclusions concerning its origin and gene flow.     

Mechanism of recruitment of WASP to the immunological synapse and of its activation following TCR ligation

F-actin polymerization following engagement of the T cell receptor (TCR) is dependent on WASP and is critical for T cell activation. The link between TCR and WASP is not fully understood. In resting cells, WASP exists in a complex with WIP, which inhibits its activation by Cdc42. We show that the adaptor protein CrkL binds directly to WIP. Further, TCR ligation results in the formation of a ZAP-70-CrkL-WIP-WASP complex, which is recruited to lipid rafts and the immunological synapse. TCR engagement also causes PKCtheta-dependent phosphorylation of WIP, causing the disengagement of WASP from the WIP-WASP complex, thereby releasing it from WIP inhibition. These results suggest that the ZAP-70-CrkL-WIP pathway and PKCtheta link TCR to WASP activation.     

Modulation of human nuclear receptor LRH-1 activity by phospholipids and SHP

The human nuclear receptor liver receptor homolog 1 (hLRH-1) plays an important role in the development of breast carcinomas. This orphan receptor is efficiently downregulated by the unusual co-repressor SHP and has been thought to be ligand-independent. We present the crystal structure at a resolution of 1.9 A of the ligand-binding domain of hLRH-1 in complex with the NR box 1 motif of human SHP, which we find contacts the AF-2 region of hLRH-1 using selective structural motifs. Electron density indicates phospholipid bound within the ligand-binding pocket, which we confirm using mass spectrometry of solvent-extracted samples. We further show that pocket mutations reduce phospholipid binding and receptor activity in vivo. Our results indicate that hLRH-1's control of gene expression is mediated by phospholipid binding, and establish hLRH-1 as a novel target for compounds designed to slow breast cancer development.     

Genotypic variation and identification of QTLs for agronomic traits,using AFLP and SSR markers in RILs of sunflower (<Emphasis Type="Italic">Helianthus annuus</Emphasis> L.)

A population of 77 recombinant inbred lines (RILs) were developed through single-seed descent from a cross between PAC-2 and RHA-266. Seeds of the above-mentioned RILs and their parents were planted in the field in a randomised complete block design with two replications. Genetic control for some agronomical traits—sowing-to-flowering date, plant height, stem diameter (SD), head diameter (HD), grain weight per plant, 1,000-grain weight (TGW) and the percentage of oil in grains—were measured for RILs and their parents. Genetic variability was observed among 77 RILs for all traits studied. Transgressive segregation occurred for some traits, and the comparison between 10% of selected RILs with the best parent showed significant difference for SD and HD as well as for TGW. A set of 123 RILs from the same cross, including the 77 above-mentioned RILs and their two parents, were screened with 409 AFLP and SSR markers, and a linkage map was constructed based on 367 markers. Several QTLs associated with the studied traits were identified. The effects of each QTL are moderate, ranging from 7% to 37%, but a high percentage of phenotypic variance is explained when considering all the covariants (TR² mean around 80% in each trait). Although the detected regions need to be more precisely mapped, the information obtained should help in marker-assisted selection.     

Golgi inheritance under a block of anterograde and retrograde traffic

In mitosis, the Golgi complex is inherited following its dispersion, equal partitioning and reformation in each daughter cell. The state of Golgi membranes during mitosis is controversial, and the role of Golgi-intersecting traffic in Golgi inheritance is unclear. We have used brefeldin A (BFA) to perturb Golgi-intersecting membrane traffic at different stages of the cell cycle and followed by live cell imaging the fate of Golgi membranes in those conditions. We observed that addition of the drug on cells in prometaphase prevents mitotic Golgi dispersion. Under continuous treatment, Golgi fragments persist throughout mitosis and accumulate in a Golgi-like structure at the end of mitosis. This structure localizes at microtubule minus ends and contains all classes of Golgi markers, but is not accessible to cargo from the endoplasmic reticulum or the plasma membrane because of the continuous BFA traffic block. However, it contains preaccumulated cargo, and intermixes with the reforming Golgi upon BFA washout. This structure also forms when BFA is added during metaphase, when the Golgi is not discernible by light microscopy. Together the data indicate that independent Golgi fragments that contain all classes of Golgi markers (and that can be isolated from other organelles by blocking anterograde and retrograde Golgi-intersecting traffic) persist throughout mitosis.