Phosphatidylinositol 3-kinase and Src family kinases are required for phosphorylation and membrane recruitment of Dok-1 in c-Kit signaling

Dok-1 is an adaptor protein that is a substrate for Bcr-Abl and other tyrosine protein kinases. The presence of pleckstrin homology and phosphotyrosine binding domains as well as multiple tyrosine phosphorylation sites suggests that Dok-1 is involved in protein-protein and/or protein-lipid interactions. Here we show that stimulation of Mo7 hematopoietic cells with c-Kit ligand (KL) induces phosphatidylinositol (PI) 3-kinase-dependent tyrosine phosphorylation and membrane recruitment of Dok-1. Addition of the K-Ras membrane-targeting motif to Dok-1 generated a constitutively membrane-bound Dok-1 protein whose tyrosine phosphorylation was independent of PI 3-kinase. Membrane localization of Dok-1 was required for its ability to function as a negative regulator of cell proliferation. Additional experiments revealed that Dok-1 associated with the juxtamembrane region and C-terminal tail of c-Kit. Lyn promoted phosphorylation of c-Kit and association of c-Kit and Dok-1. Both Lyn and Tec were capable of phosphorylating Dok-1. However, the use of primary bone marrow mast cells from normal and Lyn-deficient mice demonstrated that Lyn is required for KL-dependent Dok-1 tyrosine phosphorylation. Taken together, these data indicate that activation of PI 3-kinase by KL promotes binding of the Dok pleckstrin homology domain and Dok-1 recruitment to the plasma membrane where Dok-1 is phosphorylated by Src and/or Tec family kinases.     

Ammonoid palaeobiology

This, the second in the series of virtual issues of Palaeontology, charts the development of concepts in ammonoid palaeobiology, with reference to nine selected papers published in this journal from 1965 onwards. These cover a broad range of topics: goniatite bed palaeoecology, systematics and evolution in Scaphites, flow dynamics of cephalopod shells, shell growth and differential geometry, septal function, ammonoid assemblage analysis, evolutionary trends, stratocladistics, and soft part preservation. Despite the growth and success of palaeobiological interpretations of ammonoid evolution, life, morphology and environments over the last 50 years, ammonoid research retains a great potential for the future.     

Interleukin-15 and interleukin-15R alpha SNPs and associations with muscle, bone, and predictors of the metabolic syndrome

The aims of this study were to examine associations between two SNPs in the human IL-15 gene and three SNPs in the IL-15Ralpha gene with predictors of metabolic syndrome and phenotypes in muscle, strength, and bone at baseline and in response to resistance training (RT). Subjects were Caucasians who had not performed RT in the previous year and consisted of a strength cohort (n=748), volumetric cohort (n=722), and serum cohort (n=544). Subjects completed 12 weeks of unilateral RT of the non-dominant arm, using their dominant arm as an untrained control. ANCOVA analyses revealed gender-specific associations with: (1) IL-15 SNP (rs1589241) and cholesterol (p=0.04), LDL (p=0.02), the homeostasis model assessment (HOMA; p=0.03), and BMI (p=0.002); (2) IL-15 SNP (rs1057972) and the pre- to post-training absolute difference in 1RM strength (p=0.02), BMI (p=0.008), and fasting glucose (p=0.03); (3) IL-15Ralpha SNP (rs2296135) and baseline total bone volume (p=0.04) and the pre- to post-training absolute difference in isometric strength (p=0.01); and 4) IL-15Ralpha SNP (rs2228059) and serum triglycerides (p=0.04), baseline whole muscle volume (p=0.04), baseline cortical bone volume (p=0.04), and baseline muscle quality (p=0.04). All associations were consistent in showing a potential involvement of the IL-15 pathway with muscle and bone phenotypes and predictors of metabolic syndrome.     

Interactions between aluminium and phosphorus on root surfaces and cell wall material

Summary The incorporation of aluminium into barley roots followed similar kinetics over a 24-hour incubation at either 23°C or 3°C. At either temperature most of the aluminium was recovered in the cell wall fraction of the roots. Aluminium is adsorbed by preparations of cell wall material, and once bound is not appreciably exchangeable with calcium or sodium ions. Cell wall material pretreated with aluminium is able to adsorb appreciable amounts of phosphorus. In experiments with root segments the phosphorus associated with aluminium was found to be almost completely exchangeable. A mechanism for the interaction of aluminium and phosphorus on the root surface is proposed.Supported by S.R.C. postdoctoral fellowship.     

Cellular basis of defective sperm function and its association with the genesis of reactive oxygen species by human spermatozoa

Addition of the divalent cation ionophore, A23187, to washed populations of human spermatozoa resulted in a sudden burst of production of reactive oxygen species which peaked within 3-5 min. This activity was dependent upon the presence of calcium in the external medium and was unaffected by the mitochondrial inhibitors, oligomycin, antimycin and rotenone. Studies with scavengers of reactive oxygen species revealed that, while reagents directed against singlet oxygen and the hydroxyl radical were without effect, cytochrome C reduced the response to A23187 by about 50%, suggesting that the superoxide anion radical is a major product of the activated human spermatozoon. The clinical implications of these studies stem from the considerable variation observed between individuals in the levels of reactive oxygen species produced by the spermatozoa. This variability was shown to be inversely related to the ability of the spermatozoa to exhibit sperm-oocyte fusion on exposure to A23187; defective samples exhibited a basal level of reactive oxygen species production which was 40 times that observed with normal functional cells.     

Distribution of mercury 203 in pregnant rats and their fetuses following systemic infusions with thiol-containing amino acids and glutathione during late gestation

To investigate the effect of amino acids and the tripeptide glutathione (GSH) on tissue uptake of methylmercury (MeHg) in the developing rat fetus in utero, pregnant rats were continuously infused into the external jugular vein with 0.1 mM L-cysteine, 0.1 mM L-leucine, 0.1 mM GSH or saline commencing on day 17 of gestation. This was followed at 24, 48, and 72 hours by external jugular infusion of 50 microM [203Hg]-MeHgCl administered in 1 ml over 1 hour. Pups were surgically removed from the uterus on gestational day 21. Whole body, brain, kidney, liver, and placental 203Hg radioactivity was measured by means of gamma-spectrometry. Brain 203Hg concentration in pups exposed in utero to L-cysteine was significantly higher compared with pups exposed to saline (P less than 0.05). Brain 203Hg concentration in pups exposed in utero to L-leucine and GSH was significantly depressed compared with pups exposed to saline (P less than 0.05). Kidney 203Hg concentration was not significantly changed in all treatment groups compared with controls. Liver 203Hg concentration was significantly depressed in L-leucine- and GSH-treated pups compared with controls (P less than 0.05). Placental 203Hg concentration was not affected by any treatment compared with controls. These effects occurred despite no difference in total 203Hg body burden among pups, irrespective of the treatment. In addition, infusion with L-cysteine resulted in a significant increase in 203Hg brain concentration in dams compared with controls, and 203Hg brain concentration in L-leucine- and GSH-treated dams was significantly depressed compared with controls. Thus 203Hg distribution in both adult and developing animals is altered by chronic amino acid or GSH infusions and suggests that MeHg uptake may be mediated through the formation of a cysteine-MeHg complex which is transported across the blood-brain barrier by the neutral amino acid carrier transport system.     

Faunal turnovers and trilobite morphologies in the upper Cambrian Leptoplastus Zone at Andrarum, southern Sweden

The Furongian (upper Cambrian) Leptoplastus Zone marks a time of critical changes in the evolution of olenid trilobites. This zone, unexposed at Andrarum in Skåne, southern Sweden, has been re-excavated and the sequence of faunas and sediments logged in detail. The faunal succession accords with that previously described from borehole cores by Westergård, and the subzones of L. paucisegmentatus, L. raphidophorus, L. crassicornis, L. ovatus, L. angustatus, and L. stenotus have been recognized. In the first two subzones the olenid assemblages are monospecific. At the base of the L. crassicornis Subzone more than one species is present and morphotypes with long genal spines appear for the first time. Faunal turnover is rapid, but the incoming of new species is invariably linked to an abrupt change in sedimentation, or follows an unfossiliferous interval; species either arose or migrated in after a time of environmental perturbation. Particular faunal associations are often confined to discrete sedimentary packages though some species may range through a succession of sedimentary changes. Leptoplastus crassicornis has very long genal spines, adapted for resting on the sea floor; it may have competed with the coeval, and very similar, L. angustatus. Subsequently, L. angustatus is accompanied by the stout-bodied, short-spined L. ovatus, which presumably occupied a different niche within the same environment. Leptoplastus stenotus is convergent on the much earlier L. paucisegmentatus, and likewise is found as a monospecific assemblage, presumably being adapted to a similar niche.     

Effects of high-dose soy isoflavones and equol on reproductive tissues in female cynomolgus monkeys

Soy isoflavonoids have well-established estrogenic properties in cell culture and rodent models, raising concerns that high isoflavonoid intake may promote development of uterine and breast cancers. To address this concern we evaluated the effects of high-dose isoflavonoid supplements on reproductive tissues in a postmenopausal primate model. Thirty adult female ovariectomized monkeys (Macaca fascicularis) were randomized to receive a control diet 1) alone, 2) with 509 mg/day of the soy isoflavones genistein and daidzein (IF), or 3) with 1020 mg/day of racemic equol (EQ), an isoflavan, for approximately 1 mo. Doses are expressed in aglycone units as calorically scaled human equivalents. Total serum isoflavonoid levels 4 h postfeeding were <20 nmol/L, 2570.7 nmol/L, and 6944.8 nmol/L for control, IF, and EQ groups, respectively. Equol was the predominant serum isoflavonoid in both IF (72.5%) and EQ (99.7%) groups. Aglycones represented 0.9% (IF) and 0.5% (EQ) of total serum isoflavonoids. Histologically, uteri and mammary glands were diffusely atrophic in all groups. Uterine weight, endometrial thickness, glandular area, and epithelial proliferation in the uterus were not significantly different among treatment groups (ANOVA P > 0.1 for all). Endometrial progesterone receptor gene expression was significantly increased in the IF group (P = 0.02), while protein expression was not altered (ANOVA P > 0.1). Within the mammary gland, proliferation and indicators of estrogen exposure did not differ among treatment groups (ANOVA P > 0.1 for all). These findings indicate that high doses of dietary soy isoflavonoids have minimal uterotrophic or mammotrophic effects in an established primate model.