Receptive fields of disparity-tuned simple cells in macaque V1

Binocular simple cells in primary visual cortex (V1) are the first cells along the mammalian visual pathway to receive input from both eyes. Two models of how binocular simple cells could extract disparity information have been put forward. The phase-shift model proposes that the receptive fields in the two eyes have different subunit organizations, while the position-shift model proposes that they have different overall locations. In five fixating macaque monkeys, we recorded from 30 disparity-tuned simple cells that showed selectivity to the disparity in a random dot stereogram. High-resolution maps of the left and right eye receptive fields indicated that both phase and position shifts were common. Single cells usually showed a combination of the two, and the optimum disparity was best correlated with the sum of receptive field phase and position shift.     

Alpha-melanocyte-stimulating hormone inhibits allergic airway inflammation

Alpha-melanocyte-stimulating hormone (alpha-MSH) is a neuropeptide controlling melanogenesis in pigmentary cells. In addition, its potent immunomodulatory and immunosuppressive activity has been recently described in cutaneous inflammatory disorders. Whether alpha-MSH is also produced in the lung and might play a role in the pathogenesis of inflammatory lung conditions, including allergic bronchial asthma, is unknown. Production and functional role of alpha-MSH were investigated in a murine model of allergic airway inflammation. alpha-MSH production was detected in bronchoalveolar lavage fluids. Although aerosol challenges stimulate alpha-MSH production in nonsensitized mice, this rapid and marked stimulation was absent in allergic animals. Treatment of allergic mice with alpha-MSH resulted in suppression of airway inflammation. These effects were mediated via IL-10 production, because IL-10 knockout mice were resistant to alpha-MSH treatment. This study provides evidence for a novel function of alpha-MSH linking neuroimmune functions in allergic airway inflammation.     

Critical role of preconceptional immunization for protective and nonpathological specific immunity in murine neonates

Expression of Th2 immunity against environmental Ags is the hallmark of the allergic phenotype and contrasts with the Th1-like pattern, which is stably expressed in healthy adults throughout life. Epidemiological studies indicate that the prenatal environment plays an important and decisive role in the development of allergy later in life. Since the underlying mechanisms were unclear, an animal model was developed to study the impact of maternal allergy on the development of an allergic immune response in early life. An allergic Th2 response was induced in pregnant mice by sensitization and aerosol allergen exposure. Both, IgG1 and IgG2a, but not IgE, Abs cross the placental barrier. Free allergen also crosses the placental area and was detected in serum and amniotic fluids of neonatal F(1) mice. These F(1) mice demonstrated a suppressed Th1 response, as reflected by lowered frequencies and reduced levels of IFN-gamma production. Development of an IgE response against the same allergen was completely prevented early in life. This effect was mediated by diaplacental transfer of allergen-specific IgG1 Abs. In contrast, allergic sensitization against a different allergen early in life was accelerated in these mice. This effect was mediated by maternal CD4 and OVA-specific Th2 cells induced by allergic sensitization during pregnancy. These data indicate a critical role for maternal T and B cell response in shaping pre- and postnatal maturation of specific immunity to allergens.     

Fructose-1,6-bisphosphatase from<Emphasis Type="Italic"> Corynebacterium glutamicum</Emphasis>: expression and deletion of the<Emphasis Type="Italic"> fbp</Emphasis> gene and biochemical characterization of the enzyme

The class II fructose-1,6-bisphosphatase gene of Corynebacterium glutamicum, fbp, was cloned and expressed with a N-terminal His-tag in Escherichia coli. Purified, His-tagged fructose-1,6-bisphosphatase from C. glutamicum was shown to be tetrameric, with a molecular mass of about 140 kDa for the homotetramer. The enzyme displayed Michaelis-Menten kinetics for the substrate fructose 1,6-bisphosphate with a K_m value of about 14 µM and a V_max of about 5.4 µmol min^–1 mg^–1 and k_cat of about 3.2 s^–1. Fructose-1,6-bisphosphatase activity was dependent on the divalent cations Mg²⁺ or Mn²⁺ and was inhibited by the monovalent cation Li⁺ with an inhibition constant of 140 µM. Fructose 6-phosphate, glycerol 3-phosphate, ribulose 1,5-bisphosphate and myo-inositol-monophosphate were not significant substrates of fructose-1,6-bisphosphatase from C. glutamicum. The enzymatic activity was inhibited by AMP and phosphoenolpyruvate and to a lesser extent by phosphate, fructose 6-phosphate, fructose 2,6-bisphosphate, and UDP. Fructose-1,6-bisphosphatase activities and protein levels varied little with respect to the carbon source. Deletion of the chromosomal fbp gene led to the absence of any detectable fructose-1,6-bisphosphatase activity in crude extracts of C. glutamicum WTfbp and to an inability of this strain to grow on the carbon sources acetate, citrate, glutamate, and lactate. Thus, fbp is essential for growth on gluconeogenic carbon sources and likely codes for the only fructose-1,6-bisphosphatase in C. glutamicum.     

An ikaros-containing chromatin-remodeling complex in adult-type erythroid cells

We have previously described a SWI/SNF-related protein complex (PYR complex) that is restricted to definitive (adult-type) hematopoietic cells and that specifically binds DNA sequences containing long stretches of pyrimidines. Deletion of an intergenic DNA-binding site for this complex from a human beta-globin locus construct results in delayed human gamma- to beta-globin switching in transgenic mice, suggesting that the PYR complex acts to facilitate the switch. We now show that PYR complex DNA-binding activity also copurifies with subunits of a second type of chromatin-remodeling complex, nucleosome-remodeling deacetylase (NuRD), that has been shown to have both nucleosome-remodeling and histone deacetylase activities. Gel supershift assays using antibodies to the ATPase-helicase subunit of the NuRD complex, Mi-2 (CHD4), confirm that Mi-2 is a component of the PYR complex. In addition, we show that the hematopoietic cell-restricted zinc finger protein Ikaros copurifies with PYR complex DNA-binding activity and that antibodies to Ikaros also supershift the complex. We also show that NuRD and SWI/SNF components coimmunopurify with each other as well as with Ikaros. Competition gel shift experiments using partially purified PYR complex and recombinant Ikaros protein indicate that Ikaros functions as a DNA-binding subunit of the PYR complex. Our results suggest that Ikaros targets two types of chromatin-remodeling factors-activators (SWI/SNF) and repressors (NuRD)-in a single complex (PYR complex) to the beta-globin locus in adult erythroid cells. At the time of the switch from fetal to adult globin production, the PYR complex is assembled and may function to repress gamma-globin gene expression and facilitate gamma- to beta-globin switching.     

Automated generation of a dihydropyrimidine compound library using microwave-assisted processing

Here we report the generation of a small focused library of 12 diversely functionalized dihydropyrimidine (DHPM) derivatives via one-pot three-component Biginelli cyclocondensation of beta-ketoesters, aldehydes and (thio)ureas. By applying controlled microwave heating under sealed vessel conditions using a fully automated microwave instrument including a gripper and liquid handler, the sequential synthesis of DHPMs can be performed in a shorter reaction time (10-20 min per one DHPM derivative) compared to conventional heating methods, which normally require several hours of reflux heating. The solid products either crystallize directly upon cooling or can be precipitated upon addition of water, requiring only filtration for isolation. In this way, the DHPM derivatives are obtained in high purity and no further purification by recrystallization or chromatography is necessary. This can be ascribed to the microwave heating technology where less side-product formation is often seen. The preparation of this 12-membered DHPM library can be carried out within approximately 9 h.