Synthesis and evaluation of potential affinity labels derived from endomorphin-2.

In an attempt to identify potential peptide-based affinity labels for opioid receptors, endomorphin-2 (Tyr-Pro-Phe-PheNH2), a potent and selective endogenous ligand for mu-opioid receptors, was chosen as the parent peptide for modification. The tetrapeptide analogs were prepared using standard Fmoc-solid phase peptide synthesis in conjunction with incorporation of Fmoc-Phe(p-NHAlloc) and modification of the p-amino group. The electrophilic groups isothiocyanate and bromoacetamide were introduced into the para position on either Phe3 or Phe4; the corresponding free amine-containing peptides were also prepared for comparison. The peptides bearing an affinity label group and their free amine analogs were evaluated in a radioligand-binding assay using Chinese hamster ovary (CHO) cells expressing mu- and delta-opioid receptors. Modification on Phe4 was better tolerated than on Phe3 for mu-receptor binding. Among the analogs tested, [Phe(p-NH2)4]endomorphin-2 showed the highest affinity (IC50 = 37 nm) for mu-receptors. The Phe(p-NHCOCH2Br)4 analog displayed the highest mu-receptor affinity (IC50 = 158 nm) among the peptides containing an affinity label group. Most of the compounds exhibited negligible binding affinity for delta-receptors, similar to the parent peptide.     

Preliminary analysis of a hybrid zone between two subspecies of Zootermopsis nevadensis

For species largely allopatric in distribution, zones of contact provide an opportunity for hybridization, testing grounds for species boundaries, and may result in the formation of a new species. Thus, hybrid zones have the potential to provide important insights into speciation. In this study, we performed a preliminary analysis of a hybrid zone between two subspecies of the dampwood termite, Zootermopsis nevadensis (Z. n. nuttingi and Z. n. nevadensis) near Bartle, CA, using 12 microsatellite loci and a mitochondrial gene. Fifty-seven colonies collected in 36 locations were analyzed. The analysis of genetic markers revealed a large hybrid zone approximately 104 km in width. Although stepped clines best explained the data, we are unable to rule out the existence of a mosaic hybrid zone. We inferred weak selection (s* < 0.05%) against hybrids, but the data also suggested the existence of a barrier to gene flow from Z. n. nevadensis to Z. n. nuttingi, but not in the other direction. Given the large zone of contact, extensive sampling is needed to obtain a more complete characterization of the hybrid zone. However, the results of this study suggest that despite the accumulation of phenotypic and genetic differences and time since divergence (~2 million years), Z. n. nuttingi and Z. n. nevadensis are capable of extensive hybridization.     

Hunter-gatherer males are more risk-seeking than females,even in late childhood

Observed economic and labor disparities between the sexes may, in part, result from evolved sex differences in risk preferences. Using incentivized economic games, we report on sex differences in risk preferences in the Hadza, a population of hunter-gatherers. One game played in 2010 (n = 233) found that more Hadza males than females prefer to gamble for a chance to earn more maize rather than settle for a sure, but smaller, amount. Similarly, a second game played in 2013 (n = 102) found that male Hadza gamble a greater proportion of honey for a chance to earn more compared to female Hadza. Effect sizes are small to medium. We find weak evidence that risk-taking increases in men as their mating opportunities increase. In both games, the sex difference widens throughout childhood and is greatest among adolescents; though note that child samples are small. We explore developmental trends further using observational data on food returns in children (n = 357). Our data suggest that while the mean number of calories boys bring to camp remains stable with age, the variance in their caloric returns increases. Among girls, the variance remains stable with increased age. Both the economic games and food return data are consistent with the sexual division of labor wherein boys, beginning in late childhood, begin to target riskier foods. To the extent that the Hadza allow us to make inferences about long-standing patterns of human behavior, we suggest that sex differences in risk preferences may have been present long before agriculture and the modern work environment.     

Serine-rich repeat proteins and pili promote Streptococcus agalactiae colonization of the vaginal tract

Streptococcus agalactiae (group B streptococcus [GBS]) is a Gram-positive bacterium found in the female rectovaginal tract and is capable of producing severe disease in susceptible hosts, including newborns and pregnant women. The vaginal tract is considered a major reservoir for GBS, and maternal vaginal colonization poses a significant risk to the newborn; however, little is known about the specific bacterial factors that promote GBS colonization and persistence in the female reproductive tract. We have developed in vitro models of GBS interaction with the human female cervicovaginal tract using human vaginal and cervical epithelial cell lines. Analysis of isogenic mutant GBS strains deficient in cell surface organelles such as pili and serine-rich repeat (Srr) proteins shows that these factors contribute to host cell attachment. As Srr proteins are heavily glycosylated, we confirmed that carbohydrate moieties contribute to the effective interaction of Srr-1 with vaginal epithelial cells. Antibody inhibition assays identified keratin 4 as a possible host receptor for Srr-1. Our findings were further substantiated in an in vivo mouse model of GBS vaginal colonization, where mice inoculated with an Srr-1-deficient mutant exhibited decreased GBS vaginal persistence compared to those inoculated with the wild-type (WT) parental strain. Furthermore, competition experiments in mice showed that WT GBS exhibited a significant survival advantage over the ΔpilA or Δsrr-1 mutant in the vaginal tract. Our results suggest that these GBS surface proteins contribute to vaginal colonization and may offer new insights into the mechanisms of vaginal niche establishment.     

TPX2 regulates the localization and activity of Eg5 in the mammalian mitotic spindle

Mitotic spindle assembly requires the regulated activity of numerous spindle-associated proteins. In mammalian cells, the Kinesin-5 motor Eg5 interacts with the spindle assembly factor TPX2, but how this interaction contributes to spindle formation and function is not established. Using bacterial artificial chromosome technology, we generated cells expressing TPX2 lacking the Eg5 interaction domain. Spindles in these cells were highly disorganized with multiple spindle poles. The TPX2-Eg5 interaction was required for kinetochore fiber formation and contributed to Eg5 localization to spindle microtubules but not spindle poles. Microinjection of the Eg5-binding domain of TPX2 resulted in spindle elongation, indicating that the interaction of Eg5 with TPX2 reduces motor activity. Consistent with this possibility, we found that TPX2 reduced the velocity of Eg5-dependent microtubule gliding, inhibited microtubule sliding, and resulted in the accumulation of motor on microtubules. These results establish a novel function of TPX2 in regulating the location and activity of the mitotic motor Eg5.     

Oxidized lipid depresses canine growth,immune function,and bone formation

Dietary oxidized lipids can increase oxidative stress and potentially contribute to a variety of disease syndromes. This research describes the first use of a canine model to assess the effects of dietary oxidized lipids on growth, antioxidant status, and some immune functions. Three groups of eight, two-month old coon-hound puppies were pair fed diets for 16 weeks. The control diet contained <50 ppm aldehydes, and two additional diets contained thermally oxidized lipids targeted to contain 100 ppm aldehydes (medium-oxidation) and 500 ppm aldehydes (high-oxidation). Dogs fed the high-oxidation diet weighed less than those from the medium-oxidation (P < 0.05) and control groups (P < 0.001) at the end of the study. Oxidized lipids reduced serum vitamin E levels, total body fat content, and bone appositional rate. At different time points of the study, peripheral blood neutrophils and monocytes from dogs fed the HO diet had reduced oxidative burst capacity and produced less superoxide and hydrogen peroxide when stimulated with phorbol esters compared to the control group. Lymphocyte blastogenesis in response to concanavalin A was suppressed by dietary oxidized lipid. This study indicates that dietary oxidized lipids negatively affect the growth, antioxidant status, and some immune functions of dogs. Importantly, some effects are evident at 100 ppm aldehydes in the diet, which is a moderate level of oxidation. The rapid growth and weight gain of the dog during the first 6 months of life may also provide a better model for assessing the risks of dietary oxidized lipid in children and adolescents than previously used rodent models.