Sequential Subterranean Transport of Excavated Sand and Foraged Seeds in Nests of the Harvester Ant,Pogonomyrmex badius

During their approximately annual nest relocations, Florida harvester ants (Pogonomyrmex badius) excavate large and architecturally-distinct subterranean nests. Aspects of this process were studied by planting a harvester ant colony in the field in a soil column composed of layers of 12 different colors of sand. Quantifying the colors of excavated sand dumped on the surface by the ants revealed the progress of nest deepening to 2 m and enlargement to 8 L in volume. Most of the excavation was completed within about 2 weeks, but the nest was doubled in volume after a winter lull. After 7 months, we excavated the nest and mapped its structure, revealing colored sand deposited in non-host colored layers, especially in the upper 30 to 40 cm of the nest. In all, about 2.5% of the excavated sediment was deposited below ground, a fact of importance to sediment dating by optically-stimulated luminescence (OSL). Upward transport of excavated sand is carried out in stages, probably by different groups of ants, through deposition, re-transport, incorporation into the nest walls and floors and remobilization from these. This results in considerable mixing of sand from different depths, as indicated in the multiple sand colors even within single sand pellets brought to the surface. Just as sand is transported upward by stages, incoming seeds are transported downward to seed chambers. Foragers collect seeds and deposit them only in the topmost nest chambers from which a separate group of workers rapidly transports them downward in increments detectable as a "wave" of seeds that eventually ends in the seed chambers, 20 to 80 cm below the surface. The upward and downward transport is an example of task-partitioning in a series-parallel organization of work carried out by a highly redundant work force in which each worker usually completes only part of a multi-step process.     

Zinc-mediated inhibition of cyclic nucleotide phosphodiesterase activity and expression suppresses TNF-alpha and IL-1 beta production in monocytes by elevation of guanosine 3',5'-cyclic monophosphate

The trace element zinc affects several aspects of immune function, such as the release of proinflammatory cytokines from monocytes. We investigated the role of cyclic nucleotide signaling in zinc inhibition of LPS-induced TNF-alpha and IL-1beta release from primary human monocytes and the monocytic cell line Mono Mac1. Zinc reversibly inhibited enzyme activity of phosphodiesterase-1 (PDE-1), PDE-3, and PDE-4 in cellular lysate. It additionally reduced mRNA expression of PDE-1C, PDE-4A, and PDE-4B in intact cells. Although these PDE can also hydrolyze cAMP, only the cellular level of cGMP was increased after incubation with zinc, whereas cAMP was found to be even slightly reduced due to inhibition of its synthesis. To investigate whether an increase in cGMP alone is sufficient to inhibit cytokine release, the cGMP analogues 8-bromo-cGMP and dibutyryl cGMP as well as the NO donor S-nitrosocysteine were used. All three treatments inhibited TNF-alpha and IL-1beta release after stimulation with LPS. Inhibition of soluble guanylate cyclase-mediated cGMP synthesis with LY83583 reversed the inhibitory effect of zinc on LPS-induced cytokine release. In conclusion, inhibition of PDE by zinc abrogates the LPS-induced release of TNF-alpha and IL-1beta by increasing intracellular cGMP levels.     

The antitumor activity of hydrophobin SC3, a fungal protein

The use of mushroom extracts has been common practice in traditional medicine for centuries, including the treatment of cancer. Proteins called hydrophobins are very abundant in mushrooms. Here, it was examined whether they have antitumor activity. Hydrophobin SC3 of Schizophyllum commune was injected daily intraperitoneally starting 1 day after tumor induction in two tumor mouse models (sarcoma and melanoma). SC3 reduced the size and weight of the melanoma significantly, but the sarcoma seemed not affected. However, microscopic analysis of the tumors 12 days after induction revealed a strong antitumor effect of SC3 on both tumors. The mitotic activity of the tumor decreased 1.6- (melanoma) to 2.3-fold (sarcoma), while the vital mass decreased 2.3- (melanoma) to 4.3-fold (sarcoma) compared to the control. Treatment did not cause any signs of toxicity. Behavior, animal growth, and weight of organs were similar to animals injected with vehicle, and no histological abnormalities were found in the organs. In vitro cell culture studies revealed no direct cytotoxic effect of SC3 towards sarcoma cells, while cytotoxic activity was observed towards melanoma cells at a high SC3 concentration. Daily treatment with SC3 did not result in detectable levels of anti-SC3 antibodies in the plasma. Instead, a cellular immune response was observed. Incubation of spleen cells with SC3 resulted in a 1.5- to 2.5-fold increase in interleukin-10 and TNF-α mRNA levels. In conclusion, the nontoxic fungal hydrophobin SC3 showed tumor-suppressive activity possibly via immunomodulation and may be of benefit as adjuvant in combination with chemotherapy and radiation.     

Sec-Mediated Transport of Posttranslationally Dehydrated Peptides in Lactococcus lactis

Nisin is a lanthionine-containing antimicrobial peptide produced by Lactococcus lactis. Its (methyl)lanthionines are introduced by two posttranslational enzymatic steps involving the dehydratase NisB, which dehydrates serine and threonine residues, and the cyclase NisC, which couples these dehydrated residues to cysteines, yielding thioether-bridged amino acids called lanthionines. The prenisin is subsequently exported by the ABC transporter NisT and extracellularly processed by the peptidase NisP. L. lactis expressing the nisBTC genes can modify and secrete a wide range of nonlantibiotic peptides. Here we demonstrate that in the absence of NisT and NisC, the Sec pathway of L. lactis can be exploited for the secretion of dehydrated variants of therapeutic peptides. Furthermore, posttranslational modifications by NisB and NisC still occur even when the nisin leader is preceded by a Sec signal peptide or a Tat signal peptide 27 or 44 amino acids long, respectively. However, transport of fully modified prenisin via the Sec pathway is impaired. The extent of NisB-mediated dehydration could be improved by raising the intracellular concentration NisB or by modulating the export efficiency through altering the signal sequence. These data demonstrate that besides the traditional lantibiotic transporter NisT, the Sec pathway with an established broad substrate range can be utilized for the improved export of lantibiotic enzyme-modified (poly)peptides.     

Trifluoperazine and chlorpromazine block secretion from human platelets evoked at basal cytoplasmic free calcium by activators of C-kinase

Trifluoperazine, chlorpromazine and other drugs to inhibit calmodulin-dependent processes are also known to inhibit protein kinase C. The effect of these agents on secretion evoked by known activators of C-kinase has been studied in human platelets loaded with the fluorescent Ca indicator, quin2 and preincubated with aspirin. The secretory response stimulated by phorbol ester and exogenous diacyglycerol, at basal levels of cytoplasmic free Ca²⁺, [Ca²⁺]_i, was suppressed by trifluoperazine, chlorpromazine and W-7, as was the secretion evoked by collagen that occurs without a change in [Ca²⁺]_i, The response to thrombin, which is accompanied by elevated [Ca²⁺]_i was barely affected. Modest elevation of [Ca²⁺]_i by Ca ionophore was able to overcome the inhibitory effect of these drugs on the response to phorbol ester, diacylglycerol, and collagen.     

The rabbit alpha-like globin gene cluster is polymorphic both in the sizes of BamHI fragments and in the numbers of duplicated sets of genes

The alpha-like globin gene cluster in rabbits contains embryonic zeta- globin genes, an adult alpha-globin gene, and theta-globin genes of undetermined function. The basic arrangement of genes, deduced from analysis of cloned DNA fragments, is 5'-zeta 0-zeta 1-alpha 1-theta 1- zeta 2-zeta 3-theta 2-3'. However, the pattern of restriction fragments containing zeta- and theta-globin genes varies among individual rabbits. Analysis of BamHI fragments of genomic DNA from 24 New Zealand white rabbits revealed eight different patterns of fragments containing zeta-globin genes. The large BamHI fragments containing genes zeta 0 and zeta 1 are polymorphic in length, whereas a 1.9-kb fragment containing the zeta 2 gene and the 3.5-kb fragment containing the zeta 3 gene do not vary in size. In contrast to this constancy in the size of the restriction fragments, the copy number of the zeta 2 and zeta 3 genes does vary among different rabbits. No length polymorphism was detected in the BamHI fragments containing the theta-globin genes, but again the copy number varies for restriction fragments containing the theta 2 gene. The alpha 1- and theta 1-globin genes are located in a nonpolymorphic 7.2-kb BamHI fragment. The combined data from hybridization with both zeta and theta probes shows that the BamHI cleavage pattern does not vary within the region 5'-alpha 1-theta 1- zeta 2-zeta 3-theta 2-3', but the pattern genomic blot-hybridization patterns for the progeny of parental rabbits with different zeta-globin gene patterns shows that the polymorphic patterns are inherited in a Mendelian fashion. Two different haplotypes have been mapped based on the genomic blot-hybridization data. The variation in the alpha-like globin gene cluster in the rabbit population results both from differences in the copy number of the duplication block containing the zeta-zeta-theta gene set and from the presence or absence of polymorphic BamHI sites.      

Thrombin and ionomycin can raise platelet cytosolic Ca2+ to micromolar levels by discharge of internal Ca2+ stores: studies using fura-2

In the presence of 1 mM EGTA, the addition of the calcium ionophore ionomycin to human platelets loaded with 30 microM fura-2 could elevate [Ca2+]i from less than 100 nM to a maximum of greater than 3 microM, presumably by discharge of Ca2+ from internal stores. Under the same conditions thrombin could maximally increase [Ca2+]i to a peak of greater than 1 microM which then declined to near resting levels within 3-4 minutes; by contrast in platelets loaded with 1 mM quin2 thrombin could raise [Ca2+]i to only about 200 nM. In the presence of 1 mM Ca2+ the peak response to thrombin in fura-2-loaded platelets was higher (1.4 microM) than that observed in the presence of EGTA (1.1 microM) and the elevation in [Ca2+] was prolonged, presumably by Ca2+ influx. These results with fura-2-loaded platelets indicate that mobilisation of internal Ca2+ can contribute a substantial proportion of the early peak [Ca2+]i evoked by thrombin directly confirming the deductions from previous work with different loadings of quin2. Under natural conditions the major role of Ca2+ influx may be to prolong the [Ca2+]i rise rather than to make it larger.     

Rearranged gene order between pig and human in a QTL region on SSC 7

On porcine Chromosome 7, the region surrounding the MHC region contains QTL influencing many traits including growth, back fat thickness, and carcass composition. Towards the identification of the responsible gene(s), this article describes an increase of density of the radiated hybrid map of SSC 7 in the q11-q14 region and the comparative analysis of gene order on the porcine RH map and human genome assembly. Adding 24 new genes in this region, we were able to build a framework map that fills in gaps on the previous maps. The new software Carthagene was used to build a robust framework in this region. Comparative analysis of human and porcine maps revealed a global conservation of gene order and of distances between genes. A rearranged fragment of around 3.7 Mb was, however, found in the pig approximately 20 Mb upstream from the expected location on the basis of the human map. This rearrangement, found by RH mapping on the IMpRH 7.000 rads panel, has been confirmed by two-color FISH and by mapping on the high resolution IMNpRH2 12.000 rads panel. The rearranged fragment contains two microsatellites found at the most likely QTL location in the INRA QTL experiment. It also contains the BMP5 gene, which, together with CLPS, could be considered as a possible candidate.     

Cloning of two tryptophan hydroxylase genes expressed in the diencephalon of the developing zebrafish brain

The monoamine serotonin (5-HT) exerts key neuromodulatory activities in all animal phyla, but the development and function of the serotonergic system is still incompletely understood. The zebrafish Danio rerio is an excellent model to approach this question since it is amenable to a combination of genetic, molecular and embryological studies. In order to characterize the organization of serotonergic neurons in the zebrafish we cloned two cDNAs encoding distinct forms of tryptophan hydroxylase (Tph), the rate-limiting enzyme in serotonin synthesis. We report here the pattern of expression of these two genes in relation with immunoreactive TH and 5-HT nuclei in the developing zebrafish embryo and early larva. tphD1 expression starts at 22 h post-fertilization (hpf) in the epiphysis and in basal spinal cells. Expression persists in the epiphysis until at least 4 days (dpf). Between 48 hpf and 3 dpf, tphD1 expression is initiated in retinal amacrine cells and in restricted preoptic and posterior tubercular nuclei within the basal diencephalon. At 3 and 4 dpf, tphD1 expression is newly initiated in the caudal hypothalamus and in branchial arches-associated neurons. tphD2 mRNA is detected transiently (between 30 somites and 32 hpf) in a restricted preoptic nucleus. All sites of tphD1 or D2 expression within the anterior central nervous system are also immunoreactive for 5-HT, but are not positive for TH. However, neither tphD gene is expressed in raphe nuclei, suggesting that additional tph gene(s) exist in zebrafish to account for 5-HT synthesis in that location. The co-expression of tphD1, tphD2 and 5-HT in the zebrafish diencephalon appears in striking contrast to the situation in mammals, where diencephalic serotonin results from re-uptake rather than from local production.     

High Energy Digestion Efficiency and Altered Lipid Metabolism Contribute to Obesity in BFMI Mice

To constitute a valuable resource to identify individual genes involved in the development of obesity, a novel mouse model, the Berlin Fat Mouse Inbred line 860 (BFMI860), was established. In order to characterize energy intake and energy expenditure in obese BFMI860 mice, we performed two independent sets of experiments in male BFMI860 and B6 control mice (10 per line). In experiment 1, we analyzed body fat content noninvasively by dual‐energy X‐ray absorptiometry and measured resting metabolic rate at thermoneutrality (RMRt) and respiratory quotient (RQ) in week 6, 10, and 18. In a second experiment, energy digested (energy intake minus fecal energy loss) was determined by bomb calorimetry from week 6 through week 12. BFMI860 mice were heavier and had higher fat mass (final body fat content was 24.7% compared with 14.6% in B6). They also showed fatty liver syndrome. High body fat accumulation in BFMI860 mice was restricted to weeks 6–10 and was accompanied by hyperphagia, higher energy digestion, higher RQs, and abnormally high blood triglyceride levels. Lean mass–adjusted RMRt was not altered between lines. These results indicate that in BFMI860 mice, the excessive accumulation of body fat is associated with altered lipid metabolism, high energy intake, and energy digestion. Assuming that BFMI860 mice and their obese phenotypes are of polygenic nature, this line is an excellent model for the study of obesity in humans, especially for juvenile obesity and hyperlipidemia.