Experiment on wound activated production of Prostaglandin E2 and related toxic compounds in populations of Gracilaria vermiculophylla

The capacity of the East Asian seaweed Gracilaria vermiculophylla ("Ogonori") for production of prostaglandin E2 from arachidonic acid occasionally causes food poisoning after ingestion. During the last two decades the alga has been introduced to Europe and North America. Non-native populations have been shown to be generally less palatable to marine herbivores than native populations. We hypothesized that the difference in palatability among populations could be due to differences in the algal content of prostaglandins. We therefore compared the capacity for wound-activated production of prostaglandins and other eicosatetraenoid oxylipins among five native populations in East Asia and seven non-native populations in Europe and NW Mexico, using a targeted metabolomics approach. In two independent experiments non-native populations exhibited a significant tendency to produce more eicosatetraenoids than native populations after acclimation to identical conditions and subsequent artificial wounding. Fourteen out of 15 eicosatetraenoids that were detected in experiment I and all 19 eicosatetraenoids that were detected in experiment II reached higher mean concentrations in non-native than in native specimens. The datasets generated in both experiments are contained in http://doi.pangaea.de/10.1594/PANGAEA.855008. Wounding of non-native specimens resulted on average in 390 % more 15-keto-PGE2, in 90 % more PGE2, in 37 % more PGA2 and in 96 % more 7,8-di-hydroxy eicosatetraenoic acid than wounding of native specimens. The dataset underlying this statement is contained in http://doi.pangaea.de/10.1594/PANGAEA.854847. Not only PGE2, but also PGA2 and dihydroxylated eicosatetraenoic acid are known to deter various biological enemies of G. vermiculophylla that cause tissue or cell wounding, and in the present study the latter two compounds also repelled the mesograzer Littorina brevicula. The dataset underlying this statement is contained in http://doi.pangaea.de/10.1594/PANGAEA.854922. Non-native populations of G. vermiculophylla are thus more defended against herbivory than native populations. This increased capacity for activated chemical defense may have contributed to their invasion success and at the same time it poses an elevated risk for human food safety.

The activation domain of the enhancer binding protein p45NF-E2 interacts with TAFII130 and mediates long-range activation of the α- and β-globin gene loci in an erythroid cell line

We have used the interaction between the erythroid-specific enhancer in hypersensitivity site 2 of the human β-globin locus control region and the globin gene promoters as a paradigm to examine the mechanisms governing promoter/enhancer interactions in this locus. We have demonstrated that enhancer-dependent activation of the globin promoters is dependent on the presence of both a TATA box in the proximal promoter and the binding site for the erythroid-specific heteromeric transcription factor NF-E2 in the enhancer. Mutational analysis of the transcriptionally active component of NF-E2, p45NF-E2, localizes the critical region for this function to a proline-rich transcriptional activation domain in the NH2-terminal 80 amino acids of the protein. In contrast to the wild-type protein, expression of p45 NF-E2 lacking this activation domain in an NF-E2 null cell line fails to support enhancer-dependent transcription in transient assays. More significantly, the mutated protein also fails to reactivate expression of the endogenous β- or α-globin loci in this cell line. Protein-protein interaction studies reveal that this domain of p45 NF-E2 binds specifically to a component of the transcription initiation complex, TATA binding protein associated factor TAFII130. These findings suggest one potential mechanism for direct recruitment of distal regulatory regions of the globin loci to the individual promoters.

Parkin functions as an E2-dependent ubiquitin– protein ligase and promotes the degradation of the synaptic vesicle-associated protein, CDCrel-1

Parkinson's disease is a common neurodegenerative disorder in which familial-linked genes have provided novel insights into the pathogenesis of this disorder. Mutations in Parkin, a ring-finger-containing protein of unknown function, are implicated in the pathogenesis of autosomal recessive familial Parkinson's disease. Here, we show that Parkin binds to the E2 ubiquitin-conjugating human enzyme 8 (UbcH8) through its C-terminal ring-finger. Parkin has ubiquitin–protein ligase activity in the presence of UbcH8. Parkin also ubiquitinates itself and promotes its own degradation. We also identify and show that the synaptic vesicle-associated protein, CDCrel-1, interacts with Parkin through its ring-finger domains. Furthermore, Parkin ubiquitinates and promotes the degradation of CDCrel-1. Familial-linked mutations disrupt the ubiquitin–protein ligase function of Parkin and impair Parkin and CDCrel-1 degradation. These results suggest that Parkin functions as an E3 ubiquitin–protein ligase through its ring domains and that it may control protein levels via ubiquitination. The loss of Parkin's ubiquitin–protein ligase function in familial-linked mutations suggests that this may be the cause of familial autosomal recessive Parkinson's disease.

Growth differentiation factor-9 stimulates progesterone synthesis in granulosa cells via a prostaglandin E2/EP2 receptor pathway

Growth differentiation factor-9 (GDF-9), an oocyte-secreted member of the transforming growth factor β superfamily, progesterone receptor, cyclooxygenase 2 (Cox2; Ptgs2), and the EP2 prostaglandin E2 (PGE2) receptor (EP2; Ptgerep2) are required for fertility in female but not male mice. To define the interrelationship of these factors, we used a preovulatory granulosa cell culture system in which we added recombinant GDF-9, prostaglandins, prostaglandin receptor agonists, or cyclooxygenase inhibitors. GDF-9 stimulated Cox2 mRNA within 2 h, and PGE2 within 6 h; however, progesterone was not increased until 12 h after addition of GDF-9. This suggested that Cox2 is a direct downstream target of GDF-9 but that progesterone synthesis required an intermediate. To determine whether prostaglandin synthesis was required for progesterone production, we analyzed the effects of PGE2 and cyclooxygenase inhibitors on this process. PGE2 can stimulate progesterone synthesis by itself, although less effectively than GDF-9 (3-fold vs. 6-fold increase over 24 h, respectively). Furthermore, indomethacin or NS-398, inhibitors of Cox2, block basal and GDF-9-stimulated progesterone synthesis. However, addition of PGE2 to cultures containing both GDF-9 and NS-398 overrides the NS-398 block in progesterone synthesis. To further define the PGE2-dependent pathway, we show that butaprost, a specific EP2 agonist, stimulates progesterone synthesis and overrides the NS-398 block. In addition, GDF-9 stimulates EP2 mRNA synthesis by a prostaglandin- and progesterone-independent pathway. Thus, GDF-9 induces an EP2 signal transduction pathway which appears to be required for progesterone synthesis in cumulus granulosa cells. These studies further demonstrate the importance of oocyte–somatic cell interactions in female reproduction.