Selective binding of synapse-associated protein 97 to GluR-A alpha-amino-5-hydroxy-3-methyl-4-isoxazole propionate receptor subunit is determined by a novel sequence motif.

A family of four closely related PDZ domain-containing membrane-associated guanylate kinase homologues (MAGUKs) is involved in the regulation of the amount and functional state of ionotropic glutamate receptors in excitatory synapses. To understand the mechanisms that determine the specificity of these interactions, we examined the structural basis of the highly selective association between the ionotropic GluR subunit GluR-A and synapse-associated protein 97 (SAP97). The C terminus of GluR-A bound to the PDZ domains of SAP97, but not to those of three related MAGUKs, PSD-93, PSD-95, and SAP102. Experiments with single PDZ domains indicated that the strongest contribution was by the second PDZ domain. Unexpectedly, mutation analysis of the GluR-A C terminus revealed that a tripeptide sequence SSG at position −9 to −11 plays an essential role in this binding, in addition to a C-terminal type I PDZ binding motif (leucine at C terminus and threonine at the −2 position). Analysis of the in vitroMAGUK-binding properties of a GluR-D mutant with a one-residue deletion at the C terminus provides further support for the view that an SSG sequence located N-terminally from a type I PDZ binding motif can mediate selective binding to SAP97 and suggest the existence of a novel variation of the PDZ domain-peptide interaction

Dynamic visualization of membrane-inserted fraction of pHluorin-tagged channels using repetitive acidification technique.

Background Changes in neuronal excitability, synaptic efficacy and generally in cell signaling often result from insertion of key molecules into plasma membrane (PM). Many of the techniques used for monitoring PM insertion lack either spatial or temporal resolution. Results We improved the imaging method based on time-lapse total internal reflection fluorescence (TIRF) microscopy and pHluorin tagging by supplementing it with a repetitive extracellular acidification protocol. We illustrate the applicability of this method by showing that brief activation of NMDA receptors ("chemical LTP") in cultured hippocampal neurons induced a persistent PM insertion of glutamate receptors containing the pHluorin-tagged GluR-A(flip) subunits. Conclusion The repetitive acidification technique provides a more accurate way of monitoring the PM-inserted fraction of fluorescently tagged molecules and offers a good temporal and spatial resolution

Developmental switch in the kinase dependency of long-term potentiation depends on expression of GluA4 subunit-containing AMPA receptors

Peer reviewe

Cysteine 893 is a target of regulatory thiol modifications of GluA1 AMPA receptors

Recent studies indicate that glutamatergic signaling involves, and is regulated by, thiol modifying and redox-active compounds. In this study, we examined the role of a reactive cysteine residue, Cys-893, in the cytosolic C-terminal tail of GluA1 AMPA receptor as a potential regulatory target. Elimination of the thiol function by substitution of serine for Cys-893 led to increased steady-state expression level and strongly reduced interaction with SAP97, a major cytosolic interaction partner of GluA1 C-terminus. Moreover, we found that of the three cysteine residues in GluA1 C-terminal tail, Cys-893 is the predominant target for Snitrosylation induced by exogenous nitric oxide donors in cultured cells and lysates. Co-precipitation experiments provided evidence for native association of SAP97 with neuronal nitric oxide synthase (nNOS) and for the potential coupling of Ca2+- permeable GluA1 receptors with nNOS via SAP97. Our results show that Cys-893 can serve as a molecular target for regulatory thiol modifications of GluA1 receptors, including the effects of nitric oxide.Peer reviewe

Dynamic visualization of membrane-inserted fraction of pHluorin-tagged channels using repetitive acidification technique

<p>Abstract</p> <p>Background</p> <p>Changes in neuronal excitability, synaptic efficacy and generally in cell signaling often result from insertion of key molecules into plasma membrane (PM). Many of the techniques used for monitoring PM insertion lack either spatial or temporal resolution.</p> <p>Results</p> <p>We improved the imaging method based on time-lapse total internal reflection fluorescence (TIRF) microscopy and pHluorin tagging by supplementing it with a repetitive extracellular acidification protocol. We illustrate the applicability of this method by showing that brief activation of NMDA receptors ("chemical LTP") in cultured hippocampal neurons induced a persistent PM insertion of glutamate receptors containing the pHluorin-tagged GluR-A(flip) subunits.</p> <p>Conclusion</p> <p>The repetitive acidification technique provides a more accurate way of monitoring the PM-inserted fraction of fluorescently tagged molecules and offers a good temporal and spatial resolution.</p

Clustering environment of BL Lac object RGB 1745+398

The BL Lac object RGB 1745+398 lies in an environment that makes it possible to study the cluster around it more deeply than the environments of other BL Lac objects. The cluster centered on the BL Lac works as a strong gravitational lens, forming a large arc around itself. The aim of this paper is to study the environment and characteristics of this object more accurately than the environments of other BL Lac objects have been before.We measured the redshifts of galaxies in the cluster from the absorption lines in their spectra. The velocity dispersion was then obtained from the redshifts. The gravitational lensing was used for measuring the mass at the center of the cluster. The mass of the whole cluster could then be estimated using the softened isothermal sphere mass distribution. Finally, the richness of the cluster was determined by counting the number of galaxies near the BL Lac object and obtaining the galaxy-BL Lac spatial covariance function, $B_{gb}$. The redshifts of nine galaxies in the field were measured to be near the redshift of the BL Lac object, confirming the presence of a cluster. The average redshift of the cluster is 0.268, and the velocity dispersion $(470^{+190}_{-110})$ km s$^{-1}$. The mass of the cluster is M_{500}=(4^{+3}_{-2})\times10^{14} M_{\sun} which implies a rather massive cluster. The richness measurement also suggests that this is a rich cluster: the result for covariance function is $B_{gb}=(600\pm200)$ Mpc$^{1.77}$, which corresponds to Abell richness class 1 and which is consistent with the mass and velocity dispersion of the cluster.Comment: 5 pages, accepted to A&

Mitochondrial DNA variation in sudden cardiac death: a population-based study

Cardiomyopathy and cardiac conduction defects are common manifestations of mitochondrial disease. Previous studies suggest that clinically asymptomatic individuals harbouring pathogenic mitochondrial DNA (mtDNA) mutations in the cardiac muscle may have sudden cardiac death (SCD) as the first manifestation of mitochondrial disease. We investigated the contribution of pathogenic mtDNA point mutations and mtDNA haplogroups in cardiac muscle in a cohort of 280 Finnish subjects that had died from non-ischaemic SCD with the median age of death at 59 years and in 537 population controls. We did not find any common or novel pathogenic mutations, but the frequency of haplogroup H1 was higher in the SCD subjects than that in 537 population controls (odds ratio: 1.76, confidence interval 95%: 1.02–3.04). We conclude that, at the population level, pathogenic point mutations in mtDNA do not contribute to non-ischaemic SCD, but natural variation may modify the risk.</p