Pituitary adenylate cyclase activating polypeptide (PACAP) reduces food intake in mice

Pituitary adenylate cyclase activating peptide (PACAP) is a peptide that is present in the hypothalamus and other areas of the rat brain. This study demonstrates that PACAP reduces food intake after intracerebroventricular injection in food-deprived mice. Behavioral analysis suggests that this decrease in food intake is, in part, compensated for by an increase in other behaviors. Pituitary adenylate cyclase activating peptide also was demonstrated to antagonize increased food intake resulting from administration of neuropeptide Y. Thus, PACAP joins a growing list of neuropeptides involved in the central regulation of food intake.     

Chefs' opinions of restaurant portion sizes

Objectives: The objectives were to determine who establishes restaurant portion sizes and factors that influence these decisions, and to examine chefs’ opinions regarding portion size, nutrition information, and weight management. Research Methods and Procedures: A survey was distributed to chefs to obtain information about who is responsible for determining restaurant portion sizes, factors influencing restaurant portion sizes, what food portion sizes are being served in restaurants, and chefs’ opinions regarding nutrition information, health, and body weight. The final sample consisted of 300 chefs attending various culinary meetings. Results: Executive chefs were identified as being primarily responsible for establishing portion sizes served in restaurants. Factors reported to have a strong influence on restaurant portion sizes included presentation of foods, food cost, and customer expectations. While 76% of chefs thought that they served “regular” portions, the actual portions of steak and pasta they reported serving were 2 to 4 times larger than serving sizes recommended by the U.S government. Chefs indicated that they believe that the amount of food served influences how much patrons consume and that large portions are a problem for weight control, but their opinions were mixed regarding whether it is the customer's responsibility to eat an appropriate amount when served a large portion of food. Discussion: Portion size is a key determinant of energy intake, and the results from this study suggest that cultural norms and economic value strongly influence the determination of restaurant portion sizes. Strategies are needed to encourage chefs to provide and promote portions that are appropriate for customers’ energy requirements.     

Genetic bases of renal agenesis in the ACI rat: mapping of Renag1 to chromosome 14

Unilateral renal agenesis (URA) is a common developmental defect in humans, occurring at a frequency of approximately 1 in 500–1000 births. Several genetic syndromes include bilateral or unilateral renal agenesis as an associated phenotype. However, URA frequently occurs in individuals not afflicted by these syndromes and is often asymptomatic. Although it is clear that genetic factors contribute to the etiology of URA, the genetic bases of URA are poorly defined at this time. ACI rats, both males and females, exhibit URA at an incidence of 5%–15%. In this article we characterize the incidence of URA in female and male F₁, F₂, and backcross (BC) progeny from reciprocal genetic crosses between the ACI strain and the unaffected Brown Norway (BN) strain. Through interval mapping analyses of 353 phenotypically defined female F₂ progeny, we mapped to rat Chromosome 14 (RNO14) a genetic locus, designated Renag1 (Renal agenesis 1), that serves as the major determinant of URA in these crosses. Further genotypic analyses of URA-affected female and male F₂ and BC progeny localized Renag1 to a 14.4-Mb interval on RNO14 bounded by markers D14Rat50 and D14Rat12. The data from these genetic studies suggest that the ACI allele of Renag1 acts in an incompletely dominant and incompletely penetrant manner to confer URA. James D. Shull and Cynthia M. Lachel authors contributed equally to this work.     

Pesticide exposure: the hormonal function of the female reproductive system disrupted?

Some pesticides may interfere with the female hormonal function, which may lead to negative effects on the reproductive system through disruption of the hormonal balance necessary for proper functioning. Previous studies primarily focused on interference with the estrogen and/or androgen receptor, but the hormonal function may be disrupted in many more ways through pesticide exposure. The aim of this review is to give an overview of the various ways in which pesticides may disrupt the hormonal function of the female reproductive system and in particular the ovarian cycle. Disruption can occur in all stages of hormonal regulation: 1. hormone synthesis; 2. hormone release and storage; 3. hormone transport and clearance; 4. hormone receptor recognition and binding; 5. hormone postreceptor activation; 6. the thyroid function; and 7. the central nervous system. These mechanisms are described for effects of pesticide exposure in vitro and on experimental animals in vivo. For the latter, potential effects of endocrine disrupting pesticides on the female reproductive system, i.e. modulation of hormone concentrations, ovarian cycle irregularities, and impaired fertility, are also reviewed. In epidemiological studies, exposure to pesticides has been associated with menstrual cycle disturbances, reduced fertility, prolonged time-to-pregnancy, spontaneous abortion, stillbirths, and developmental defects, which may or may not be due to disruption of the female hormonal function. Because pesticides comprise a large number of distinct substances with dissimilar structures and diverse toxicity, it is most likely that several of the above-mentioned mechanisms are involved in the pathophysiological pathways explaining the role of pesticide exposure in ovarian cycle disturbances, ultimately leading to fertility problems and other reproductive effects. In future research, information on the ways in which pesticides may disrupt the hormonal function as described in this review, can be used to generate specific hypotheses for studies on the effects of pesticides on the ovarian cycle, both in toxicological and epidemiological settings.     

Theoretical ab initio study of the effects of methylation on structure and stability of G:C Watson-Crick base pair

Methylation of DNA occurs most readily at N(3), N(7), and O(6) of purine bases and N(3) and O(2) of pyrimidines. Methylated bases are continuously formed through endogenous and exogenous mechanisms. The results of a theoretical ab initio study on the methylation of G:C base pair components are reported. The geometries of the local minima were optimized without symmetry restrictions by the gradient procedure at DFT level of theory and were verified by energy second derivative calculations. The standard 6-31G(d) basis set was used. The single-point calculations have been performed at the MP2/6-31G(d,p), MP2/6-31++G(d,p), and MP2/6-311++G(2d,2p) levels of theory. The geometrical parameters, relative stability and counterpoise corrected interaction energies are reported. Also, using a variation-perturbation energy decomposition scheme we have found the vital contributions to the total interaction energy.     

Closely related G-protein-coupled receptors use multiple and distinct domains on G-protein alpha-subunits for selective coupling

The molecular basis of selectivity in G-protein receptor coupling has been explored by comparing the abilities of G-protein heterotrimers containing chimeric Galpha subunits, comprised of various regions of Gi1alpha, Gtalpha, and Gqalpha, to stabilize the high affinity agonist binding state of serotonin, adenosine, and muscarinic receptors. The data indicate that multiple and distinct determinants of selectivity exist for individual receptors. While the A1 adenosine receptor does not distinguish between Gi1alpha and Gtalpha sequences, the 5-HT1A and 5-HT1B serotonin and M2 muscarinic receptors can couple with Gi1 but not Gt. It is possible to distinguish domains that eliminate coupling and are defined as "critical," from those that impair coupling and are defined as "important." Domains within the N terminus, alpha4-helix, and alpha4-helix-alpha4/beta6-loop of Gi1alpha are involved in 5-HT and M2 receptor interactions. Chimeric Gi1alpha/Gqalpha subunits verify the critical role of the Galpha C terminus in receptor coupling, however, the individual receptors differ in the C-terminal amino acids required for coupling. Furthermore, the EC50 for interactions with Gi1 differ among the individual receptors. These results suggest that coupling selectivity ultimately involves subtle and cooperative interactions among various domains on both the G-protein and the associated receptor as well as the G-protein concentration.     

Association of the Ste20-like kinase (SLK) with the microtubule. Role in Rac1-mediated regulation of actin dynamics during cell adhesion and spreading

Cytoskeletal remodeling events are tightly regulated by signal transduction systems that impinge on adhesion components and modulators of cellular architecture. We have previously shown that the Ste20-like kinase (SLK) can induce apoptosis through the induction of actin disassembly and cellular retraction (Sabourin, L. A., Tamai, K., Seale, P., Wagner, J., and Rudnicki, M. A. (2000) Mol. Cell. Biol. 20, 684-696). Using immunofluorescence studies, we report that SLK is redistributed with adhesion components at large podosome-like adhesion sites in fibronectin-stimulated fibroblasts. However, in vitro kinase assays demonstrate that its activity is not modulated following fibronectin stimulation. Double immunofluorescence studies in exponentially growing or spreading fibroblasts show that SLK is associated with the microtubule network and can be coprecipitated with alpha-tubulin. Furthermore, the stimulation of adhesion site formation by microtubule-disrupting agents induces the relocalization of SLK with unpolymerized alpha-tubulin to large vinculin-containing adhesion complexes. Using microinjection studies, we show that ectopic expression of activated SLK induces the disassembly of actin stress fibers, a process that can be inhibited by dominant negative Rac1. Significantly, endogenous SLK can be colocalized with Rac1 in spreading cells on FN. Finally, the overexpression of SLK by adenoviral infection inhibits cell spreading on fibronectin. These results suggest that SLK is part of a microtubule-associated complex that is targeted to adhesion sites and implicated in the regulation of cytoskeletal dynamics.     

Transformation of Saccharopolyspora erythraea by electroporation of germinating spores: construction of propionyl Co-A carboxylase mutants

The introduction of plasmid DNA into germinating spores of an industrially improved strain of Saccharopolyspora erythraea was accomplished by electroporation. Various parameters affecting the efficiency of electroporation were examined. The most critical factor was the extent of spore germination. Electrocompetence was limited to a 4-h period following the initial emergence of the germ tube. Electroporation efficiencies as high as 2 × 10⁵ CFU μg⁻¹ of plasmid DNA were obtained using electrocompetent germlings. The optimal field strength was 12–14 kV cm⁻¹ with a pulse duration of 15–20 ms. Electrocompetent germlings were stored at −80°C without a significant decrease in transformation efficiency. The utility of this protocol was demonstrated by isolating a propionyl-CoA carboxylase mutant through targeted gene disruption and replacement. Received 3 April 1998/ Accepted in revised form 28 September 1998     

Sequence variations in small-subunit ribosomal RNAs of Hartmannella vermiformis and their phylogenetic implications

Evidence of associations between free-living amoebas and human disease has been increasing in recent years. Knowledge about phylogenetic relationships that may be important for the understanding of pathogenicity in the genera involved is very limited at present. Consequently, we have begun to study these relationships and report here on the phylogeny of Hartmannella vermiformis, a free-living amoeba that can harbor the etiologic agent of Legionnaires' disease. Our analysis is based on studies of small-subunit ribosomal RNA genes (srDNA). Nucleotide sequences were determined for nuclear srDNA from three strains of H. vermiformis isolated from the United Kingdom, Germany, and the United States. These sequences then were compared with a sequence previously obtained for a North American isolate by J. H. Gunderson and M. L. Sogin. The four genes are 1,840 bp long, with an average GC content of 49.6%. Sequence differences among the strains range are 0.38%-0.76%. Variation occurs at 19 positions and includes 2 single-base indels plus 14 monotypic and 3 ditypic single-base substitutions. Variation is limited to eight helix/loop structures according to a current model for srRNA secondary structure. Parsimony, distance, and bootstrap analyses used to examine phylogenetic relationships between the srDNA sequences of H. vermiformis and other eukaryotes indicated that Hartmannella sequences were most closely related to those of Acanthamoeba and the alga Cryptomonas. All ditypic sites were consistent with a separation between European and North American strains of Hartmannella, but results of other tests of this relationship were statistically inconclusive.