Protein Kinase A Opposes the Phosphorylation-dependent Recruitment of Glycogen Synthase Kinase 3β to A-kinase Anchoring Protein 220

The proximity of an enzyme to its substrate can influence rate and magnitude of catalysis. A-kinase anchoring protein 220 (AKAP220) is a multivalent anchoring protein that can sequester a variety of signal transduction enzymes. These include protein kinase A (PKA) and glycogen synthase kinase 3β (GSK3β). Using a combination of molecular and cellular approaches we show that GSK3β phosphorylation of Thr-1132 on AKAP220 initiates recruitment of this kinase into the enzyme scaffold. We also find that AKAP220 anchors GSK3β and its substrate β-catenin in membrane ruffles. Interestingly, GSK3β can be released from the multienzyme complex in response to PKA phosphorylation on serine 9, which suppresses GSK3β activity. The signaling scaffold may enhance this regulatory mechanism, as AKAP220 has the capacity to anchor two PKA holoenzymes. Site 1 on AKAP220 (residues 610–623) preferentially interacts with RII, whereas site 2 (residues 1633–1646) exhibits a dual specificity for RI and RII. In vitro affinity measurements revealed that site 2 on AKAP220 binds RII with ∼10-fold higher affinity than site 1. Occupancy of both R subunit binding sites on AKAP220 could provide a mechanism to amplify local cAMP responses and enable cross-talk between PKA and GSK3β.     

Interindividual influence on diurnal rhythms of activity in cycling and noncycling populations of the field vole,Microtus agrestis L.

Summary The effects of interaction among individuals with respect to wheelrunning activity has been investigated in two geographically separated populations of the field vole Microtus agrestis L. In one of them, a northern cyclically varying population, a strictly nocturnal activity pattern is changed into a more or less short term 24 h pattern under conditions of increased contact among individuals. In the other population, south-Swedish, without population cycles, no such effects were observed. This indicates population differences in response to interindividual contact which might be of importance to the known differences in population dynamics.     

Exposure of cell surface structures inEscherichia coli as studied with ultrastructural immunohistochemistry

The surfaces of two smoothEscherichia coli O6 strains, one with K13 capsular antigen and one without demonstrable capsule, and one roughE. coli strain were analyzed by using a sandwich technique with ultrastructural immunohistochemistry. Peroxidase-conjugated anti-rabbit serum and antisera corresponding to the distal lipopolysaccharide ofE. coli (O antigen), to the capsule (K antigen), or to the lipid part of the lipopolysaccharide, lipid A, were used. Using the anti-capsule serum on the smooth-capsule-producing strain, we observed the capsule as loosely connected clusters covering less of the surface than the O antigen. No reactions were observed with the capsule-negative strain having the same O antigen. The O antigen was identified all over the outer membrane in a patchy pattern with knobs covering the whole surface on the capsule-negative strain. Fewer knobs, but more variable in size than on the capsule-negative strain, were observed on the capsule-producing strain. The rough strain showed no reactions with either the O or K antiserum. Lipid A was not available for anti-lipid A serum on the rough or smooth strains.     

Cytoplasmic Ca2+ concentration of single normal human and bovine parathyroid cells measured by dual wavelength microfluorometry

Dual wavelength microfluorometry was utilized to measure the cytoplasmic calcium concentration (Ca_i ²⁺) of single parathyroid cells loaded with the indicator fura-2. The method enabled the first registrations of Ca_i ²⁺ of normal human parathyroid cells, available only in minute numbers. At 0.5 mM extracellular Ca²⁺, the Ca_i ²⁺ levels were similar in normal human and bovine cells. Both cell types responded with an initial Ca_i ²⁺ transient followed by a sustained increase when raising extracellular Ca²⁺ to 3.0 mM. The sustained effect exhibited a sigmoidal relation to extracellular Ca²⁺ in the 0.5–3.0 mM range. Although the increase was somewhat greater in the human cells, the half maximal responses were obtained at almost identical extracellular Ca²⁺ concentrations. Whereas K⁺ depolarization decreased Ca_i ²⁺, the Ca_i ²⁺ channel blocker D-600 had dual actions, raising Ca_i ²⁺at 0.5 mM Ca²⁺ and decreasing it at 3.0 mM Ca_i ²⁺, and the effects were similar in the bovine and human cells. The present experimental approach verified the validity of utilizing bovine cells as controls in studies of human parathyroid tissue and it appears suitable for analysis of the role of different subpopulations of parathyroid cells in the abnormal parathyroid tissue of patients with hyperparathyroidism.     

Chimaeras and the origins of the holopelagic annelids Typhloscolecidae and Lopadorhynchidae: a reply to Struck & Halanych (2010)

Nygren, A. & Fredrik, P. (2010). Chimaeras and the origins of the holopelagic annelids Typhloscolecidae and Lopadorhynchidae: a reply to Struck & Halanych. —Zoologica Scripta, 40, 112–114.     

External diffusion in solid-phase immunoassays

Calculations are presented describing the influence of external diffusion in the kinetics of solid-phase immunoassays. The analysis is concerned with systems where one reactant is immobilized at the surface of a sphere of arbitrary radius. The solution for a plane surface is found as a limiting case. The factors determining whether the reaction is diffusion or reaction controlled are found to be sphere radius, surface concentration of binding sites, forward reaction rate and diffusion constant of reacting species. Means of determining whether the reaction is diffusion or reaction controlled from observable quantities are described. When applied to heterogeneous antibody-antigen binding it is found that normally the binding to cell-size spheres is not limited by external diffusion. However, when applied to solid-phase assays with high surface concentrations of binding sites immobilized at plane surfaces or macroscopic spheres the binding is found to be diffusion limited. The importance of a mass transfer analysis in this case is also discussed.     

Effect of hyperthyroidism on spontaneous physical activity and energy expenditure in rats.

Thyroid hormone excess is associated with increased energy expenditure. The contributions of increases in spontaneous physical activity and nonexercise activity thermogenesis (NEAT) to this effect have not been defined. To address the hypothesis that hyperthyroidism is associated with increased spontaneous physical activity and NEAT, we rendered rats hyperthyroid by using continuous infusion of high-dose triiodothyronine for 14 days and measured the effects on physical activity and NEAT. On day 14, in the hyperthyroid group the mean +/- SD triiodothyronine concentration was 755 +/- 137 (range 574-919) ng/dl and in the control group 59 +/- 0.5 (58-59) ng/dl. Over the 14-day treatment period, mean spontaneous physical activity increased in the hyperthyroid rats from 24 +/- 7 to 36 +/- 6 activity units (AU)/min; P < 0.001 but did not increase in the controls (23 +/- 7 vs. 22 +/- 4 AU/min). Also, over the 14-day period, daily NEAT increased in the hyperthyroid rats from 8.1 +/- 2.8 to 19.7 +/- 5.0 kcal/day (P < 0.001) but did not increase in the controls (8.7 +/- 3.5 cf 9.4 +/- 1.7 kcal/day; not significant). In conclusion, hyperthyroidism is associated with increased spontaneous physical activity and NEAT.     

Genetic relationships between Nereimyra punctata and N. woodsholea (Hesionidae, Polychaeta)

We present a COI-based parsimony analysis of the relationships between the shallow water, pigmented hesionid polychaete Nereimyra punctata , and a deep-water, unpigmented form with sympatric distribution in Norway and Sweden. Apart from the pigmentation differences, the two forms exhibit no observed morphological differences. The terminals are represented by four specimens each of the two forms from the Trondheimsfjord in Norway, and four each of the two forms from northern Bohuslän in Sweden, plus members of the two hesionids Heteropodarke and Ophiodromus as outgroups. In addition, the analysis includes a topotype of the morphologically similar and unpigmented Nereimyra woodsholea from the Middle Atlantic Bight off the US east coast. The equally weighted matrix includes 132 informative characters. All most-parsimonious trees unequivocally indicate that specimens belonging to the same form (pigmented or unpigmented) from different areas are cladistically closer related to each other than different forms from the same areas. Nereimyra woodsholea is nested within the unpigmented deeper group of the Norwegian and Swedish specimens, thus indicating that this name should be applied to the deep-water form in Norway and Sweden.