The Smc5/Smc6/MAGE Complex Confers Resistance to Caffeine and Genotoxic Stress in Drosophila melanogaster

The SMC5/6 protein complex consists of the Smc5, Smc6 and Non-Smc-Element (Nse) proteins and is important for genome stability in many species. To identify novel components in the DNA repair pathway, we carried out a genetic screen to identify mutations that confer reduced resistance to the genotoxic effects of caffeine, which inhibits the ATM and ATR DNA damage response proteins. This approach identified inactivating mutations in CG5524 and MAGE, homologs of genes encoding Smc6 and Nse3 in yeasts. The fact that Smc5 mutants are also caffeine-sensitive and that Mage physically interacts with Drosophila homologs of Nse proteins suggests that the structure of the Smc5/6 complex is conserved in Drosophila. Although Smc5/6 proteins are required for viability in S. cerevisiae, they are not essential under normal circumstances in Drosophila. However, flies carrying mutations in Smc5, Smc6 and MAGE are hypersensitive to genotoxic agents such as ionizing radiation, camptothecin, hydroxyurea and MMS, consistent with the Smc5/6 complex serving a conserved role in genome stability. We also show that mutant flies are not compromised for pre-mitotic cell cycle checkpoint responses. Rather, caffeine-induced apoptosis in these mutants is exacerbated by inhibition of ATM or ATR checkpoint kinases but suppressed by Rad51 depletion, suggesting a functional interaction involving homologous DNA repair pathways that deserves further scrutiny. Our insights into the SMC5/6 complex provide new challenges for understanding the role of this enigmatic chromatin factor in multi-cellular organisms.     

EB1 Levels Are Elevated in Ascorbic Acid (AA)-stimulated Osteoblasts and Mediate Cell-Cell Adhesion-induced Osteoblast Differentiation

Osteoblasts are differentiated mesenchymal cells that function as the major bone-producing cells of the body. Differentiation cues including ascorbic acid (AA) stimulation provoke intracellular changes in osteoblasts leading to the synthesis of the organic portion of the bone, which includes collagen type I α1, proteoglycans, and matrix proteins, such as osteocalcin. During our microarray analysis of AA-stimulated osteoblasts, we observed a significant up-regulation of the microtubule (MT) plus-end binding protein, EB1, compared with undifferentiated osteoblasts. EB1 knockdown significantly impaired AA-induced osteoblast differentiation, as detected by reduced expression of osteoblast differentiation marker genes. Intracellular examination of AA-stimulated osteoblasts treated with EB1 siRNA revealed a reduction in MT stability with a concomitant loss of β-catenin distribution at the cell cortex and within the nucleus. Diminished β-catenin levels in EB1 siRNA-treated osteoblasts paralleled an increase in phospho-β-catenin and active glycogen synthase kinase 3β, a kinase known to target β-catenin to the proteasome. EB1 siRNA treatment also reduced the expression of the β-catenin gene targets, cyclin D1 and Runx2. Live immunofluorescent imaging of differentiated osteoblasts revealed a cortical association of EB1-mcherry with β-catenin-GFP. Immunoprecipitation analysis confirmed an interaction between EB1 and β-catenin. We also determined that cell-cell contacts and cortically associated EB1/β-catenin interactions are necessary for osteoblast differentiation. Finally, using functional blocking antibodies, we identified E-cadherin as a major contributor to the cell-cell contact-induced osteoblast differentiation.     

Brief communication: Evolution of a specific O allele (O1vG542A) supports unique ancestry of Native Americans

In this study, we explore the geographic and temporal distribution of a unique variant of the O blood group allele called O1v^G542A, which has been shown to be shared among Native Americans but is rare in other populations. O1v^G542A was previously reported in Native American populations in Mesoamerica and South America, and has been proposed as an ancestry informative marker. We investigated whether this allele is also found in the Tlingit and Haida, two contemporary indigenous populations from Alaska, and a pre‐Columbian population from California. If O1v^G542A is present in Na‐Dene speakers (i.e., Tlingits), it would indicate that Na‐Dene speaking groups share close ancestry with other Native American groups and support a Beringian origin of the allele, consistent with the Beringian Incubation Model. If O1v^G542A is found in pre‐Columbian populations, it would further support a Beringian origin of the allele, rather than a more recent introduction of the allele into the Americas via gene flow from one or more populations which have admixed with Native Americans over the past five centuries. We identified this allele in one Na‐Dene population at a frequency of 0.11, and one ancient California population at a frequency of 0.20. Our results support a Beringian origin of O1v^G542A, which is distributed today among all Native American groups that have been genotyped in appreciable numbers at this locus. This result is consistent with the hypothesis that Na‐Dene and other Native American populations primarily derive their ancestry from a single source population. Am J Phys Anthropol 151:649–657, 2013. © 2013 Wiley Periodicals, Inc.     

Exploring prehistory in the North American southwest with mitochondrial DNA diversity exhibited by Yumans and Athapaskans

A recent study of mitochondrial DNA variation in Native American populations from the American Southwest detected signatures of a population expansion of subhaplogroup B2a, dated to 2,105 years before present (99.5% confidence interval, 1,273–3,773 YBP), following the introduction and intensification of maize agriculture in the region. Only one Yuman group and no Athapaskan speakers were analyzed in previous studies. Here we report mtDNA haplogroup and hypervariable region (HVR I, and II) sequence data from 263 extant Yuman speakers, representing the major branches of the Yuman language family, in addition to the Western Apache (Athapaskan) to further investigate the demographic context and geographic extent of this expansion. Data presented indicate that the expansion of B2a is only slightly older [2,410 YBP (99.5% CI: 1,458–4,320 YBP)] than previously estimated and not significantly. Despite large confidence intervals there are implications for the origin and expansion of the Yuman language family. Cultural transformations due to the inundation and draining of Lake Cahuilla may explain in part the frequencies of this lineage among the Kumeyaay and other Yuman and Takic groups in Southern California. This may have been the result of group fissions and fusions followed by migration and interaction that included expanded trade networks and intermarriage among Yuman speakers. In addition, a series of in‐situ genetic bottlenecks is proposed to have occurred among the Western Apache leading to increasing homogeneity within haplogroup A, culminating in an admixture event with the Yavapai. Am J Phys Anthropol 150:618–631, 2013. © 2013 Wiley Periodicals, Inc.     

Effects of Diet Composition on Postprandial Energy Availability during Weight Loss Maintenance

Background

The major circulating metabolic fuels regulate hunger, and each is affected by dietary composition. An integrated measure of postprandial energy availability from circulating metabolic fuels may help inform dietary recommendations for weight maintenance after weight loss.

Aim

We examined the effect of low-fat (LF, 60% of energy from carbohydrate, 20% fat, 20% protein), low-glycemic index (LGI, 40%–40%-20%), and very low-carbohydrate (VLC, 10%–60%-30%) diets on total postprandial metabolic fuel energy availability (EA) during weight loss maintenance.

Methods

Eight obese young adults were fed a standard hypocaloric diet to produce 10–15% weight loss. They were then provided isocaloric LF, LGI, and VLC diets in a randomized crossover design, each for a 4-week period of weight loss maintenance. At the end of each dietary period, a test meal representing the respective diet was provided, and blood samples were obtained every 30 minutes for 5 hours. The primary outcome was EA, defined as the combined energy density (circulating level×relative energy content) of glucose, free fatty acids, and β-hydroxybutyrate. Secondary outcomes were individual metabolic fuels, metabolic rate, insulin, glucagon, cortisol, epinephrine, and hunger ratings. Respiratory quotient was a process measure. Data were analyzed by repeated-measures analysis of variance, with outcomes compared in the early (30 to 150 min) and late (180 to 300 min) postprandial periods.

Results

EA did not differ between the test meals during the early postprandial period (p = 0.99). However, EA in the late postprandial period was significantly lower after the LF test meal than the LGI (p<0.0001) and VLC (p<0.0001) test meals. Metabolic rate also differed in the late postprandial period (p = 0.0074), with higher values on the VLC than LF (p = 0.0064) and LGI (p = 0.0066) diets.

Conclusion

These findings suggest that an LF diet may adversely affect postprandial EA and risk for weight regain during weight loss maintenance.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00315354","term_id":"NCT00315354"}}NCT00315354     

B Cell Deficient Mice Are Protected from Biliary Obstruction in the Rotavirus-Induced Mouse Model of Biliary Atresia

A leading theory regarding the pathogenesis of biliary atresia (BA) is that bile duct injury is initiated by a virus infection, followed by an autoimmune response targeting bile ducts. In experimental models of autoimmune diseases, B cells have been shown to play an important role. The aim of this study was to determine the role of B cells in the development of biliary obstruction in the Rhesus rotavirus (RRV)-induced mouse model of BA. Wild-type (WT) and B cell-deficient (Ig-α^-/-) mice received RRV shortly after birth. Ig-α^-/- RRV-infected mice had significantly increased disease-free survival rate compared to WT RRV-infected BA mice (76.8% vs. 17.5%). In stark contrast to the RRV-infected BA mice, the RRV-infected Ig-α^-/- mice did not have hyperbilirubinemia or bile duct obstruction. The RRV-infected Ig-α^-/- mice had significantly less liver inflammation and Th1 cytokine production compared to RRV-infected WT mice. In addition, Ig-α^-/- mice had significantly increased numbers of regulatory T cells (Tregs) at baseline and after RRV infection compared to WT mice. However, depletion of Tregs in Ig-α^-/- mice did not induce biliary obstruction, indicating that the expanded Tregs in the Ig-α^-/- mice were not the sole reason for protection from disease. Conclusion: B cell deficient Ig-α^-/- mice are protected from biliary obstruction in the RRV-induced mouse model of BA, indicating a primary role of B cells in mediating disease pathology. The mechanism of protection may involve lack of B cell antigen presentation, which impairs T-cell activation and Th1 inflammation. Immune modulators that inhibit B cell function may be a new strategy for treatment of BA.     

Innovative levers for sustainable integration of gender medicine into medical school curricula

Efforts to integrate gender medicine into medical school curricula have focused largely on the work of individual champions. Online sex and gender materials for undergraduate courses have also been developed and disseminated. Success has been sporadic, with varying uptake across schools within and between countries. International trends in medical school accreditation processes and the growing force of the millennial student voice offer untapped opportunities to promote more systematic integration of gender medicine on a national and international level. In this commentary, the president and CEO of the Association of Faculties of Medicine of Canada and the Scientific Director of the Institute of Gender and Health of the Canadian Institutes of Health Research jointly reflect on top-down and bottom-up levers for sustainable innovation in gender medicine for undergraduate medical training.