Perilipin ablation results in a lean mouse with aberrant adipocyte lipolysis, enhanced leptin production, and resistance to diet-induced obesity

Perilipin coats the lipid droplets of adipocytes and is thought to have a role in regulating triacylglycerol hydrolysis. To study the role of perilipin in vivo, we have created a perilipin knockout mouse. Perilipin null (peri−/−) and wild-type (peri+/+) mice consume equal amounts of food, but the adipose tissue mass in the null animals is reduced to ≈30% of that in wild-type animals. Isolated adipocytes of perilipin null mice exhibit elevated basal lipolysis because of the loss of the protective function of perilipin. They also exhibit dramatically attenuated stimulated lipolytic activity, indicating that perilipin is required for maximal lipolytic activity. Plasma leptin concentrations in null animals were greater than expected for the reduced adipose mass. The peri−/− animals have a greater lean body mass and increased metabolic rate but they also show an increased tendency to develop glucose intolerance and peripheral insulin resistance. When fed a high-fat diet, the perilipin null animals are resistant to diet-induced obesity but not to glucose intolerance. The data reveal a major role for perilipin in adipose lipid metabolism and suggest perilipin as a potential target for attacking problems associated with obesity.

Pseudomonas syringae Hrp type III secretion system and effector proteins

Pseudomonas syringae is a member of an important group of Gram-negative bacterial pathogens of plants and animals that depend on a type III secretion system to inject virulence effector proteins into host cells. In P. syringae, hrp/hrc genes encode the Hrp (type III secretion) system, and avirulence (avr) and Hrp-dependent outer protein (hop) genes encode effector proteins. The hrp/hrc genes of P. syringae pv syringae 61, P. syringae pv syringae B728a, and P. syringae pv tomato DC3000 are flanked by an exchangeable effector locus and a conserved effector locus in a tripartite mosaic Hrp pathogenicity island (Pai) that is linked to a tRNALeu gene found also in Pseudomonas aeruginosa but without linkage to Hrp system genes. Cosmid pHIR11 carries a portion of the strain 61 Hrp pathogenicity island that is sufficient to direct Escherichia coli and Pseudomonas fluorescens to inject HopPsyA into tobacco cells, thereby eliciting a hypersensitive response normally triggered only by plant pathogens. Large deletions in strain DC3000 revealed that the conserved effector locus is essential for pathogenicity but the exchangeable effector locus has only a minor role in growth in tomato. P. syringae secretes HopPsyA and AvrPto in culture in a Hrp-dependent manner at pH and temperature conditions associated with pathogenesis. AvrPto is also secreted by Yersinia enterocolitica. The secretion of AvrPto depends on the first 15 codons, which are also sufficient to direct the secretion of an Npt reporter from Y. enterocolitica, indicating that a universal targeting signal is recognized by the type III secretion systems of both plant and animal pathogens.

Haploinsufficiency of Sox9 results in defective cartilage primordia and premature skeletal mineralization

In humans, SOX9 heterozygous mutations cause the severe skeletal dysmorphology syndrome campomelic dysplasia. Except for clinical descriptions, little is known about the pathogenesis of this disease. We have generated heterozygous Sox9 mutant mice that phenocopy most of the skeletal abnormalities of this syndrome. The Sox9+/− mice died perinatally with cleft palate, as well as hypoplasia and bending of many skeletal structures derived from cartilage precursors. In embryonic day (E)14.5 heterozygous embryos, bending of radius, ulna, and tibia cartilages was already prominent. In E12.5 heterozygotes, all skeletal elements visualized by using Alcian blue were smaller. In addition, the overall levels of Col2a1 RNA at E10.5 and E12.5 were lower than in wild-type embryos. We propose that the skeletal abnormalities observed at later embryonic stages were caused by delayed or defective precartilaginous condensations. Furthermore, in E18.5 embryos and in newborn heterozygotes, premature mineralization occurred in many bones, including vertebrae and some craniofacial bones. Because Sox9 is not expressed in the mineralized portion of the growth plate, this premature mineralization is very likely the consequence of allele insufficiency existing in cells of the growth plate that express Sox9. Because the hypertrophic zone of the heterozygous Sox9 mutants was larger than that of wild-type mice, we propose that Sox9 also has a role in regulating the transition to hypertrophic chondrocytes in the growth plate. Despite the severe hypoplasia of cartilages, the overall organization and cellular composition of the growth plate were otherwise normal. Our results suggest the hypothesis that two critical steps of the chondrocyte differentiation pathway are sensitive to Sox9 dosage. First, an early step presumably at the stage of mesenchymal condensation of cartilage primordia, and second, a later step preceding the transition of chondrocytes into hypertrophic chondrocytes.

Antitumor promotion by phenolic antioxidants: inhibition of AP-1 activity through induction of Fra expression.

Induction of phase 2 detoxification enzymes by phenolic antioxidants can account for prevention of tumor initiation but cannot explain why these compounds inhibit tumor promotion. Phase 2 genes are induced through an antioxidant response element (ARE). Although the ARE resembles an AP-1 binding site, we show that the major ARE binding and activating protein is not AP-1. Interestingly, AP-1 DNA binding activity was induced by the phenolic antioxidant tert-butylhydroquinone (BHQ), but the induction of AP-1 transcriptional activity by the tumor promoter 12-O-tetradecanoylphorbol 13-acetate (TPA) was inhibited by this compound. BHQ induced expression of c-jun, junB, fra-1, and fra-2, which encode AP-1 components, but was a poor inducer of c-fos and had no effect on fosB. Like c-Fos and FosB, the Fra proteins heterodimerize with Jun proteins to form stable AP-1 complexes. However, Fra-containing AP-1 complexes have low transactivation potential. Furthermore, Fra-1 repressed AP-1 activity induced by either TPA or expression of c-Jun and c-Fos. We therefore conclude that inhibitory AP-1 complexes composed of Jun-Fra heterodimers, induced by BHQ, antagonize the transcriptional effects of the tumor promoter TPA, which are mediated by Jun-Fos heterodimers. Since AP-1 is an important mediator of tumor promoter action, these findings may explain the anti-tumor-promoting activity of phenolic antioxidants.

Basis for selection of improved carbohydrate-binding single-chain antibodies from synthetic gene libraries.

A technique is described for the simultaneous and controlled random mutation of all three heavy or light chain complementarity-determining regions (CDRs) in a single-chain Fv specific for the O polysaccharide of Salmonella serogroup B. Sense oligonucleotides were synthesized such that the central bases encoding a CDR were randomized by equimolar spiking with A, G, C, and T at a level of 10% while the antisense strands contained inosine in the spiked regions. Phage display of libraries assembled from the spiked oligonucleotides by a synthetic ligase chain reaction demonstrated a bias for selection of mutants that formed dimers and higher oligomers. Kinetic analyses showed that oligomerization increased association rates in addition to slowing dissociation rates. In combination with some contribution from reduced steric clashes with residues in heavy-chain CDR2, oligomerization resulted in functional affinities that were much higher than that of the monomeric form of the wild-type single-chain Fv.

Arabidopsis COP1 protein specifically interacts in vitro with a cytoskeleton-associated protein, CIP1.

Arabidopsis COP1 acts inside the nucleus to suppress photomorphogenic cellular development, and light inactivation of COP1 may involve a specific control of its nuclear activity in hypocotyls and cotyledons, but not in roots, of developing seedlings. To understand the molecular mechanisms of COP1 action during light-mediated development, we initiated a screen for Arabidopsis cDNAs encoding proteins which interact directly with COP1 in vitro as a step to identify the cellular components involved. We report here the isolation and characterization of a cDNA clone encoding a protein designated CIP1 (COP1-interactive protein 1). CIP1 is predominantly alpha-helical and most likely involved in coiled-coil formation. It interacts specifically with the putative coiled-coil region of COP1 in vitro. Further, CIP1 is encoded by a single gene in Arabidopsis, and its mRNA and protein levels are not regulated by light. Immunofluorescent labeling of CIP1 in Arabidopsis seedling protoplasts demonstrated that CIP1 is part of, or associated with, a cytoskeletal structure in hypocotyl and cotyledon cells, but not in roots. Our results are consistent with a possible role of CIP1 in mediating light control of COP1 nuclear activity by regulating its nucleocytoplasmic partitioning.

Modularisation épistatique des loci à trait quantitatif associés à la pression artérielle et identification de gènes candidats pour l’hypertension

Problématique: L’hypertension artérielle essentielle, facteur de risque majeur dans le développement des maladies cardiovasculaires, est un trait multigénique complexe dont les connaissances sur le déterminisme génétique nécessitent d’être approfondies. De nombreux loci à trait quantitatif (QTLs); soit des gènes responsables de faire varier la pression artérielle (PA), ont été identifiés chez l’humain et le modèle animal. Cependant, le mystère plane encore sur la façon dont ces gènes fonctionnent ensemble pour réguler la PA. Hypothèse et objectif: Plutôt qu’une addition de QTLs ayant chacun une action infinitésimale sur la PA, une interaction épistatique entre les gènes serait responsable du phénotype hypertendu. Ainsi, l’étude de cette épistasie entre les gènes impliqués, directement ou indirectement, dans l’homéostasie de la PA nous permettrait d’explorer de nouvelles voies de régulation moléculaire en cause dans cette maladie. Méthodes: Via la réalisation de souches congéniques de rats, où un segment chromosomique provenant d’une souche receveuse hypertendue (Dahl Salt Sensitive, SS/Jr) est remplacé par son homologue provenant d’une souche donneuse normotendue (Lewis, LEW), des QTLs peuvent être mis en évidence. Dans ce contexte, la combinaison de QTLs via la création de doubles ou multiples congéniques constitue la première démonstration fonctionnelle des interactions intergéniques. Résultats: Vingt-sept combinaisons au total nous ont menés à l’appréciation d’une modularisation des QTLs. Ces derniers ont été catégorisés selon deux principaux modules épistatiques (EMs) où les QTLs appartenant à un même EM sont épistatiques entre eux et participent à une même voie régulatrice. Les EMs/cascades agissent alors en parallèle pour réguler la PA. Grâce à l’existence de QTLs ayant des effets opposés sur la PA, nous avons pu établir l’ordre hiérarchique entre trois paires de QTLs. Cependant, lorsque cette suite régulatrice ne peut être déterminée, d’autres approches sont nécessaires. Nos travaux nous ont mené à l’identification d’un QTL situé sur le chromosome 16 du rat (C16QTL), appartenant au EM1 et qui révélerait une nouvelle voie de l’homéostasie de la PA. Le gène retinoblastoma-associated protein 140 (Rap140)/family with sequence similarity 208 member A (Fam208a), présentant une mutation non synonyme entre SS/Jr et LEW est le gène candidat le plus plausible pour représenter C16QTL. Celui-ci code pour un facteur de transcription et semblerait influencer l’expression de Solute carrier family 7 (cationic amino acid transporter, y+ system) member 12 (Slc7a12), spécifiquement et significativement sous exprimé dans les reins de la souche congénique portant C16QTL par rapport à la souche SS/Jr. Rap140/Fam208a agirait comme un inhibiteur de la transcription de Slc7a12 menant à une diminution de la pression chez Lewis. Conclusions: L’architecture complexe de la régulation de la PA se dévoile mettant en scène de nouveaux acteurs, pour la plupart inconnus pour leur implication dans la PA. L’étude de la nouvelle voie de signalisation Rap140/Fam208a - Slc7a12 nous permettra d’approfondir nos connaissances quant à l’homéostasie de la pression artérielle et de l’hypertension chez SS/Jr. À long terme, de nouveaux traitements anti-hypertenseurs, ciblant plus d’une voie de régulation à la fois, pourraient voir le jour.

Meng xue tian wen jiao ke shu /

Chu deng xiao xue tang xue sheng yong shu.

Huang qing jing jie : [1408 juan /

On double leaves.

Meng xue du ben quan shu : 7 bian /

Xun chang xiao xue tang du shu ke sheng tu yong jiao ke shu.

Song Wenxian gong quan ji : 83 juan /

Fu: Nian pu : 3 juan / Zhu Xingti deng zuan ; Sun Jinzheng zeng -- Qian xi lu : 7 juan / Ding Lizhong bian ; Sun Jinzheng zeng bu.

Hong Beijiang shi wen ji : 66 juan /

Fu: Nian pu : 1 juan / Lü Pei deng zhuan.

Wan Jiye xian sheng qun shu yi bian : [12 juan] /

Fu zhu: You Yu yin Zhaolin, Shanyin Yu shi Zhisheng zhen cang, ji Yu shi Zhisheng jian cang shu hua yin deng zhang.

Gu jin xing shi shu bian zheng : 40 juan /

Block print.