Molecular Epidemiologic Evidence for Diabetogenic Effects of Dioxin Exposure in U.S. Air Force Veterans of the Vietnam War

BACKGROUND: One of the outcomes positively associated with dioxin exposure in humans is type 2 diabetes. OBJECTIVES: This study was conducted in order to find the molecular biological evidence for the diabetogenic action of dioxin in adipose samples from Vietnam veterans. METHODS: We obtained 313 adipose tissue samples both from Vietnam veterans who were exposed to dioxin (Operation Ranch Hand) and from comparison veterans who served in Southeast Asia with no record of dioxin exposure. We conducted quantitative reverse-transcribed polymerase chain reaction studies on selected marker mRNAs from these samples. RESULTS: We found the most sensitive and reliable molecular indicator of dioxin-induced diabetes to be the ratio of mRNA of glucose transporter 4 (GLUT4) and nuclear transcription factor kappa B (NFκB), a marker of inflammation. This ratio showed significant correlations to serum dioxin residues and to fasting glucose among those in the Ranch Hand group and, surprisingly, even in the comparison group, who have low levels of dioxin comparable to the general public. Such a correlation in the comparison group was particularly significant among those with known risk factors such as obesity and family history of diabetes. CONCLUSIONS: These results show that the GLUT4:NFκB ratio is a reliable marker for the diabetogenic action of dioxin, particularly at very low exposure levels that are not much higher than those found in the general public, implying a need to address current exposure levels

Silencing Inhibits Cre-Mediated Recombination of the Z/AP and Z/EG Reporters in Adult Cells

BACKGROUND: The Cre-loxP system has been used to enable tissue specific activation, inactivation and mutation of many genes in vivo and has thereby greatly facilitated the genetic dissection of several cellular and developmental processes. In such studies, Cre-reporter strains, which carry a Cre-activated marker gene, are frequently utilized to validate the expression profile of Cre transgenes, to act as a surrogate marker for excision of a second allele, and to irreversibly label cells for lineage tracing experiments. PRINCIPAL FINDINGS: We have studied three commonly used Cre-reporter strains, Z/AP, Z/EG and R26R-EYFP and have demonstrated that although each reporter can be reliably activated by Cre during early development, exposure to Cre in adult hematopoietic cells results in a much lower frequency of marker-positive cells in the Z/AP or Z/EG strains than in the R26R-EYFP strain. In marker negative cells derived from the Z/AP and Z/EG strains, the transgenic promoter is methylated and Cre-mediated recombination of the locus is inhibited. CONCLUSIONS: These results show that the efficiency of Cre-mediated recombination is not only dependent on the genomic context of a given loxP-flanked sequence, but also on stochastic epigenetic mechanisms underlying transgene variegation. Furthermore, our data highlights the potential shortcomings of utilizing the Z/AP and Z/EG reporters as surrogate markers of excision or in lineage tracing experiments

Quantitative Comparison of Constitutive Promoters in Human ES cells

BACKGROUND: Constitutive promoters that ensure sustained and high level gene expression are basic research tools that have a wide range of applications, including studies of human embryology and drug discovery in human embryonic stem cells (hESCs). Numerous cellular/viral promoters that ensure sustained gene expression in various cell types have been identified but systematic comparison of their activities in hESCs is still lacking. METHODOLOGY/PRINCIPAL FINDINGS: We have quantitatively compared promoter activities of five commonly used constitutive promoters, including the human β-actin promoter (ACTB), cytomegalovirus (CMV), elongation factor-1α, (EF1α), phosphoglycerate kinase (PGK) and ubiquitinC (UbC) in hESCs. Lentiviral gene transfer was used to ensure stable integration of promoter-eGFP constructs into the hESCs genome. Promoter activities were quantitatively compared in long term culture of undifferentiated hESCs and in their differentiated progenies. CONCLUSION/SIGNIFICANCE: The ACTB, EF1α and PGK promoters showed stable activities during long term culture of undifferentiated hESCs. The ACTB promoter was superior by maintaining expression in 75-80% of the cells after 50 days in culture. During embryoid body (EB) differentiation, promoter activities of all five promoters decreased. Although the EF1α promoter was downregulated in approximately 50% of the cells, it was the most stable promoter during differentiation. Gene expression analysis of differentiated eGFP+ and eGFP- cells indicate that promoter activities might be restricted to specific cell lineages, suggesting the need to carefully select optimal promoters for constitutive gene expression in differentiated hESCs

Host Shifts from Lamiales to Brassicaceae in the Sawfly Genus Athalia

Plant chemistry can be a key driver of host shifts in herbivores. Several species in the sawfly genus Athalia are important economic pests on Brassicaceae, whereas other Athalia species are specialized on Lamiales. These host plants have glucosides in common, which are sequestered by larvae. To disentangle the possible direction of host shifts in this genus, we examined the sequestration specificity and feeding deterrence of iridoid glucosides (IGs) and glucosinolates (GSs) in larvae of five species which either naturally sequester IGs from their hosts within the Plantaginaceae (Lamiales) or GSs from Brassicaceae, respectively. Furthermore, adults were tested for feeding stimulation by a neo-clerodane diterpenoid which occurs in Lamiales. Larvae of the Plantaginaceae-feeders did not sequester artificially administered p-hydroxybenzylGS and were more deterred by GSs than Brassicaceae-feeders were by IGs. In contrast, larvae of Brassicaceae-feeders were able to sequester artificially administered catalpol (IG), which points to an ancestral association with Lamiales. In line with this finding, adults of all tested species were stimulated by the neo-clerodane diterpenoid. Finally, in a phylogenetic tree inferred from genetic marker sequences of 21 Athalia species, the sister species of all remaining 20 Athalia species also turned out to be a Lamiales-feeder. Fundamental physiological pre-adaptations, such as the establishment of a glucoside transporter, and mechanisms to circumvent activation of glucosides by glucosidases are therefore necessary prerequisites for successful host shifts between Lamiales and Brassicaceae

Colonizations cause diversification of host preferences: a mechanism explaining increased generalization at range boundaries expanding under climate change

International audienceAs species' poleward range limits expand under climate change, generalists are expected to be better colonists than specialists, extending their ranges faster. This effect of specialization on range shifts has been shown, but so has the reverse cause–effect: in a global meta-analysis of butterfly diets, it was range expansions themselves that caused increases in population-level diet breadth. What could drive this unexpected process? We provide a novel behavioral mechanism by showing that, in a butterfly with extensive ecotypic variation, Edith's checkerspot, diet breadths increased after colonization events as diversification of individual host preferences pulled novel hosts into population diets. Subsequently, populations that persisted reverted toward monophagy. We draw together three lines of evidence from long-term studies of 15 independently evolving populations. First, direct observations showed a significant increase in specialization across decades: in recent censuses, eight populations used fewer host genera than in the 1980s while none used more. Second, behavioral preference-testing experiments showed that extinctions and recolonizations at two sites were followed, at first by diversification of heritable preference ranks and increases in diet breadth, and subsequently by homogenization of preferences and contractions of diet breadth. Third, we found a significant negative association in the 1980s between population-level diet breadth and genetic diversity. Populations with fewer mtDNA haplotypes had broader diets, extending to 3–4 host genera, while those with higher haplotype diversity were more specialized. We infer that diet breadth had increased in younger, recently colonized populations. Preference diversification after colonization events, whether caused by (cryptic) host shifts or by release of cryptic genetic variation after population bottlenecks, provides a mechanism for known effects of range shifts on diet specialization. Our results explain how colonizations at expanding range margins have increased population-level diet breadths, and predict that increasing specialization should accompany population persistence as current range edges become range interiors