ABHD11, a new diacylglycerol lipase involved in weight gain regulation.

Obesity epidemic continues to spread and obesity rates are increasing in the world. In addition to public health effort to reduce obesity, there is a need to better understand the underlying biology to enable more effective treatment and the discovery of new pharmacological agents. Abhydrolase domain-containing protein 11 (ABHD11) is a serine hydrolase enzyme, localized in mitochondria, that can synthesize the endocannabinoid 2-arachidonoyl glycerol (2AG) in vitro. In vivo preclinical studies demonstrated that knock-out ABHD11 mice have a similar 2AG level as WT mice and exhibit a lean metabolic phenotype. Such mice resist to weight gain in Diet Induced Obesity studies (DIO) and display normal biochemical plasma parameters. Metabolic and transcriptomic analyses on serum and tissues of ABHD11 KO mice from DIO studies show a modulation in bile salts associated with reduced fat intestinal absorption. These data suggest that modulating ABHD11 signaling pathway could be of therapeutic value for the treatment of metabolic disorders.

A Strategy Combining Differential Low-Throughput Screening and Virtual Screening (DLS-VS) Accelerating the Discovery of new Modulators for the Orphan GPR34 Receptor.

The DLS-VS strategy was developed as an integrated method for identifying chemical modulators for orphan GPCRs. It combines differential low-throughput screening (DLS) and virtual screening (VS). The two cascaded techniques offer complementary advantages and allow the experimental testing of a minimal number of compounds. First, DLS identifies modulators specific for the considered receptor among a set of receptors, through the screening of a small library with diverse chemical compounds. Then, an active molecular model of the receptor is built by homology to a validated template, and it is progressively refined by rotamers modification for key side-chains, by VS of the already screened library, and by iterative selection of the model generating the best enrichment. The refined active model is finally used for the VS of a large chemical library and the selection of a small set of compounds for experimental testing. Applied to the orphan receptor GPR34, the DLS-VS strategy combined the experimental screening of 20 000 compounds and the virtual screening of 1 250 000 compounds. It identified one agonist and eight inverse agonists, showing a high chemical diversity. We describe the method. The strategy can be applied to other GPCRs.

Identification of BCAP, a new protein associated with basal bodies and centrioles.

Cilia exert critical functions in numerous organisms, including that of cell motility, fluid transport and protozoan locomotion. Defects in this organelle can lead to lethal pathologies in humans, including primary ciliary dyskinesia. An understanding of the cilia formation process would lead to better characterization of defects involved in such pathologies. In the present study, we identified a gene encoding a novel human protein, BCAP for Basal body Centriole-Associated Protein, which shares homologies with a previously described protein, Outer Dense Fiber 2 (ODF2). ODF2, a major component of the sperm tail cytoskeleton, is required for the formation of mother centriole distal/subdistal appendages and the generation of primary cilia. Here, we show that the bcap gene contains 18 alternatively spliced exons and encodes five different isoforms, three long and two short ones. BCAP is preferentially expressed in cilia/flagella containing tissues. Moreover, its expression is correlated with cilia formation during mucociliary differentiation of human nasal epithelial cells. Using immunofluorescence analyses, BCAP was localized within basal bodies of ciliated cells and within centrioles of proliferating cells. In light of the several spliced isoforms of BCAP and the particular localization of the protein, BCAP isoforms could play distinct roles in cilia and in centrosomes.

Identification of ICIS-1, a new protein involved in cilia stability.

Cilia are specialized organelles that exert critical functions in numerous organisms, including that of cell motility, fluid transport and protozoan locomotion. Ciliary architecture and function strictly depend on basal body formation, migration and axoneme elongation. Numerous ultrastructural studies have been undertaken in different species to elucidate the process of ciliogenesis. Recent analyses have led to identification of genes specifically expressed in ciliated organisms, but most proteins involved in ciliogenesis remain uncharacterized. Using human nasal epithelial cells capable of ciliary differentiation in vitro, differential display was carried out to identify new proteins associated with ciliogenesis. We isolated a new gene, ICIS-1 (Involved in CIlia Stability-1), upregulated during mucociliary differentiation. This gene is localized within the TGF-beta1 promoter and is ubiquitously expressed in human tissues. Functional analyses of gene expression inhibition by RNA interference in Paramecium tetraurelia indicated that the ICIS-1 homologue interfered with cilia stability or formation. These findings demonstrate that ICIS-1 is a new protein associated with ciliated cells and potentially related to cilia stability.

Molecular cloning, gene localization, and structure of human cyclin B3.

Here we describe the molecular cloning of human cyclin B3, its localization, and its structure. It is localized in the subcentromeric region of the X chromosome, still not completely sequenced by the Human Genome Project. This cyclin B3 is unusually large for a mitotic cyclin. Its mRNAs were found in all tissues and were particularly abundant in testis. At least three splice variants were found in the ORF and three variants in the 5'UTR.