Identification of a GM1-Binding Protein on the Surface of Murine Neuroblastoma Cells

S20Y murine neuroblastoma cells appear to express a protein component(s) able to adhere specifically to the oligosaccharide portion of GM1 (oligo-GM1). To identify proteins with which the oligo-GM1 becomes closely associated, a radiolabeled (125I), photoactivatable derivative of oligo-GM1 was prepared. This was accomplished by reductive amination of the glucosyl moiety of oligo-GM1 to 1-deoxy-1-aminoglucitol, followed by reaction of the amine with sulfosuccinimidyl 2-(p-azidosalicylamido)ethyl-1,3'-dithiopropionate (SASD). Crosslinking studies using the photoactivatable probe indicated that it came in close proximity to a protein with an apparent molecular mass of approximately 71 kDa. In competition experiments, as little as a 10-fold molar excess of oligo-GM1 resulted in a selective reduction in labeling of this protein; preincubation with a 200-fold molar excess of siayllactose was necessary to observe the same change in the labeling pattern, lending additional support to the hypothesis that the approximately 71-kDa protein specifically associates with oligo-GM1. Cell surface location of the oligo-GM1 binding protein was confirmed using subcellular fractionation and morphological analyses.     

Regulatory T Cells Condition Lymphatic Endothelia for Enhanced Transendothelial Migration

1. Download : Download high-res image (154KB)
2. Download : Download full-size image

     

MiR-204 regulates type 1 IP3R to control vascular smooth muscle cell contractility and blood pressure

MiR-204 is expressed in vascular smooth muscle cells (VSMC). However, its role in VSMC contraction is not known. We determined if miR-204 controls VSMC contractility and blood pressure through regulation of sarcoplasmic reticulum (SR) calcium (Ca²⁺) release. Systolic blood pressure (SBP) and vasoreactivity to VSMC contractile agonists (phenylephrine (PE), thromboxane analogue (U46619), endothelin-1 (ET-1), angiotensin-II (Ang II) and norepinephrine (NE) were compared in aortas and mesenteric resistance arteries (MRA) from miR-204^−/− mice and wildtype mice (WT). There was no difference in basal systolic blood pressure (SBP) between the two genotypes; however, hypertensive response to Ang II was significantly greater in miR-204^−/− mice compared to WT mice. Aortas and MRA of miR-204^−/− mice had heightened contractility to all VSMC agonists. In silico algorithms predicted the type 1 Inositol 1, 4, 5-trisphosphate receptor (IP₃R1) as a target of miR-204. Aortas and MRA of miR-204^−/− mice had higher expression of IP₃R1 compared to WT mice. Difference in agonist-induced vasoconstriction between miR-204^−/− and WT mice was abolished with pharmacologic inhibition of IP₃R1. Furthermore, Ang II-induced aortic IP₃R1 was greater in miR-204^−/− mice compared to WT mice. In addition, difference in aortic vasoconstriction to VSMC agonists between miR-204^−/− and WT mice persisted after Ang II infusion. Inhibition of miR-204 in VSMC in vitro increased IP₃R1, and boosted SR Ca²⁺ release in response to PE, while overexpression of miR-204 downregulated IP₃R1. Finally, a sequence-specific nucleotide blocker that targets the miR-204-IP₃R1 interaction rescued miR-204-induced downregulation of IP₃R1. We conclude that miR-204 controls VSMC contractility and blood pressure through IP₃R1-dependent regulation of SR calcium release.     

Pharmacological and Physicochemical Properties of Pre-Versus Postsynaptic 5-Hydroxytryptamine1A Receptor Binding Sites in the Rat Brain: A Quantitative Autoradiographic Study

Numerous data suggested that the pharmacological and biochemical properties of 5-hydroxytryptamine1A (5-HT1A) receptors exhibit some regional differences in the CNS, notably within the raphe nuclei compared with various forebrain areas (such as the hippocampus). This possibility has been further investigated in the dorsal raphe nucleus and two areas within the hippocampus, the dentate gyrus and the CA1 area, using the quantitative autoradiographic technique. The potencies of 5'-guanylylimidodiphosphate to inhibit the specific binding of 125I-Bolton-Hunter-8-methoxy-2-(N-propyl-N-propylamino)tetralin (125I-BH-8-MeO-N-PAT) to 5-HT1A sites and of N-ethylmaleimide to block these sites irreversibly were identical in the dorsal raphe nucleus and the hippocampal areas in rat brain sections. In contrast, slight but significant differences were noted in the pH dependence and pharmacological properties of 5-HT1A sites labeled by 125I-BH-8-MeO-N-PAT in these three regions. Similarly, heat denaturation experiments and tissue exposure to either phospholipase A2 or the alkylating agent 8-methoxy-2-(N-2'-chloropropyl,N-propyl)aminotetraline revealed regional differences in the properties of 5-HT1A sites. However, in most cases, the observed variations were of greater amplitude between the CA1 area and the dentate gyrus, where 5-HT1A sites are located postsynaptically, than between any one of these areas and the dorsal raphe nucleus where they act as (presynaptic) somatodendritic autoreceptors. These data further support that subtypes of 5-HT1A receptors probably exist in the rat brain, but this heterogeneity seems unrelated to the pre- or post-synaptic location of these receptors.     

TRAIL Receptors Serve as Stress-Associated Molecular Patterns to Promote ER-Stress-Induced Inflammation

1. Download : Download high-res image (188KB)
2. Download : Download full-size image

     

Protonic and cationic changes during cyclical cation transport driven by electron transfer

A method is described for the subcellular fractionation of goldfish xanthophores. The procedure produces relatively pure fractions of caroteniod droplets, pterinosomes, cytosol and what appears to be plasma membrane. The presence of a distinct pattern of proteins is shown to be associated with the carotenoid droplets. Treatment of the xanthophores with ACTH affects the buoyant density of some carotenoid droplets and stimulates the phosphorylation of a polypeptide associated with the carotenoid droplets.     

Photoproduction of hydrogen, nitrogenase and hydrogenase activities of free and immobilized whole cells of Rhodobacter sphaeroides O.U. 001

Abstract Photoproduction of hydrogen, nitrogenase activity (acetylene reduction) and hydrogenase activity (methylene blue dye reduction) were studied in free and alginate immobilized whole cells of a purple non-sulfur photosynthetic bacterium Rhodobacter sphaeroides O.U. 001. Four-fold increase in hydrogen production, two-fold increase in nitrogenase activity and 1.2-fold increase in the hydrogenase activity were observed in immobilized cells compared to free cells. Effect of various inhibitors (CO and C₂H₂) and electron donor (H₂) on the above three functions by free and immobilized cells has also been studied.     

Assessment and comparison of neural morphology through metrical feature extraction and analysis in neuron and neuron–glia cultures

The morphology of dissociated single cerebellar Purkinje cells obtained from wild-type P1 CD1 mice was assessed in the absence and in the presence of glia. A dedicated noninvasive technique based on optical microscopy was developed. Image processing algorithms were implemented to extract metrical features characterizing cell structure and dendritic arborization. The morphological features were analyzed in order to identify quantitative differences in Purkinje cell morphology due to interactions with astrocytes.