An interaction between L-prostaglandin D synthase and arrestin increases PGD2 production

L-type prostaglandin synthase (L-PGDS) produces PGD(2), a lipid mediator involved in neuromodulation and inflammation. Here, we show that L-PGDS and arrestin-3 (Arr3) interact directly and can be co-immunoprecipitated endogenously from MG-63 osteoblasts. Perinuclear L-PGDS/Arr3 co-localization is observed in PGD(2)-producing MG-63 cells and is induced by the addition of the L-PGDS substrate or co-expression of COX-2 in HEK293 cells. Inhibition of L-PGDS activity in MG-63 cells triggers redistribution of Arr3 and L-PGDS to the cytoplasm. Perinuclear localization of L-PGDS is detected in wild-type mouse embryonic fibroblasts (MEFs) but is more diffused in MEFs-arr-2(-/-)-arr-3(-/-). Arrestin-3 promotes PGD(2) production by L-PGDS in vitro. IL-1β-induced PGD(2) production is significantly lower in MEFs-arr-2(-/-)-arr-3(-/-) than in wild-type MEFs but can be rescued by expressing Arr2 or Arr3. A peptide corresponding to amino acids 86-100 of arrestin-3 derived from its L-PGDS binding domain stimulates L-PGDS-mediated PGD(2) production in vitro and in MG-63 cells. We report the first characterization of an interactor/modulator of a PGD(2) synthase and the identification of a new function for arrestin, which may open new opportunities for improving therapies for the treatment of inflammatory diseases.     

Protein crystallography under xenon and nitrous oxide pressure: comparison with in vivo pharmacology studies and implications for the mechanism of inhaled anesthetic action

In contrast with most inhalational anesthetics, the anesthetic gases xenon (Xe) and nitrous oxide (N(2)O) act by blocking the N-methyl-d-aspartate (NMDA) receptor. Using x-ray crystallography, we examined the binding characteristics of these two gases on two soluble proteins as structural models: urate oxidase, which is a prototype of a variety of intracellular globular proteins, and annexin V, which has structural and functional characteristics that allow it to be considered as a prototype for the NMDA receptor. The structure of these proteins complexed with Xe and N(2)O were determined. One N(2)O molecule or one Xe atom binds to the same main site in both proteins. A second subsite is observed for N(2)O in each case. The gas-binding sites are always hydrophobic flexible cavities buried within the monomer. Comparison of the effects of Xe and N(2)O on urate oxidase and annexin V reveals an interesting relationship with the in vivo pharmacological effects of these gases, the ratio of the gas-binding sites' volume expansion and the ratio of the narcotic potency being similar. Given these data, we propose that alterations of cytosolic globular protein functions by general anesthetics would be responsible for the early stages of anesthesia such as amnesia and hypnosis and that additional alterations of ion-channel membrane receptor functions are required for deeper effects that progress to "surgical" anesthesia.     

High levels of tryptamine accumulation in transgenic tobacco expressing tryptophan decarboxylase

A full-length complementary DNA clone encoding tryptophan decarboxylase (TDC; EC 4.1.1.28) from Catharanthus roseus (De Luca V, Marineau C, Brisson N [1989] Proc Natl Acad Sci USA 86: 2582-2586) driven by the CaMV 35S promoter was introduced into tobacco (Nicotiana tabacum) to direct the synthesis of the protoalkaloid tryptamine from endogenous tryptophan. Young, fully expanded leaves of CaMV 35S-TDC transformed plants had from four to 45 times greater TDC activity than did controls. Tryptamine accumulated in transgenic plants to levels that were directly proportional to their TDC specific activity. Despite their increased tryptamine content, the growth and development of the CaMV 35S-TDC plants appeared normal with no significant differences in indole-3-acetic acid levels between high tryptamine and control plants. Plants with the highest TDC activity contained more than 1 milligram of tryptamine per gram fresh weight, a 260-fold increase over controls.     

In vitro embryo production in the cow: an effective alternative to the conventional embryo production approach

Development of new technology related to in vitro embryo production has allowed for the commercial use of this method of reproduction. In the present work, we evaluate the efficiency of this technology compared with conventional embryo production based on results obtained with a standard procedure, including the sexing of embryos. The donor animals were mature nonlactating dairy cows (n = 92) kept under a constant environment and feeding program in an ET center. Ultrasound guided transvaginal ovum pick-up following 48 h pre-treatment with FSH has been used for the IVF-IVC protocol. A total of 437 oocyte recovery sessions performed on 92 cows yielded 4145 oocytes, which were used in an IVF-IVC protocol. Using the conventional approach, 156 embryo collections on 49 cows yielded 1652 ova and embryos. All Quality 1 and 2 embryos were sexed by a PCR procedure, and embryos of the desired sex were transferred to synchronized recipients located at the center. The results obtained in the IVF protocol showed that 4 oocyte collections per cow performed within 60 d, yielded 38 oocytes, which resulted in 18.8 viable embryos, of which 7.05 were female. After transfer of the female embryos, an average of 3.8 recipients were pregnant at 60 d. One embryo collection under the conventional approach yielded an average of 1.2 female pregnancies, which was confirmed during the same 60-d time period. These results indicate that IVF procedures can effectively replace conventional embryo production methods when a predetermined number of pregnancies of known sex are needed within a short period of time.     

Assembly of the eastern North American herpetofauna: new evidence from lizards and frogs

Darwin first recognized the importance of episodic intercontinental dispersal in the establishment of worldwide biotic diversity. Faunal exchange across the Bering Land Bridge is a major example of such dispersal. Here, we demonstrate with mitochondrial DNA evidence that three independent dispersal events from Asia to North America are the source for almost all lizard taxa found in continental eastern North America. Two other dispersal events across Beringia account for observed diversity among North American ranid frogs, one of the most species-rich groups of frogs in eastern North America. The contribution of faunal elements from Asia via dispersal across Beringia is a dominant theme in the historical assembly of the eastern North American herpetofauna.