Biomedical subjects

J M George

Publications and source records attributed to J M George.

At least 19 recordsLinked to original sources

PubMed2003

Epitope mapping and specificity of the anti-alpha-synuclein monoclonal antibody Syn-1 in mouse brain and cultured cell lines.

While alpha- and beta-synuclein largely overlap in their expression in the vertebrate brain, only alpha-synuclein accumulates in the fibrillar aggregates typical of Parkinson's disease. It is thus critical to have immunological reagents that distinguish between these two protein isoforms. The monoclonal antibody Syn-1 (Transduction Labs) has been frequently used for the specific detection of alpha-synuclein. In this report, the epitope for Syn-1 is localized within residues 91-99 of human alpha-synuclein. Sequence differences exist in this domain that account for the specificity of Syn-1 for alpha- versus beta-synuclein. However, Syn-1 also displays reactivity with additional species (approximately 45 kDa) in brain homogenates from both wild-type and alpha-synuclein null mice, indicating a potential for cross-reactivity with a protein species that is unrelated to alpha-synuclein in brain tissue or extracts.

Animals↗

PubMed2002

Magnetism of the Fe/ZnSe(001) interface.

The magnetism of epitaxial ultrathin films of Fe on ZnSe(001) has been investigated by x-ray magnetic circular dichroism down to the submonolayer regime. In contrast to other metallic ferromagnet/semiconductor interfaces, no reduction of the Fe magnetic moment was found at the Fe/ZnSe(001) interface. Furthermore, a significant enhancement of the Fe magnetic moment compared to the bulk value was observed for coverages up to one monolayer in agreement with theoretical predictions. We also demonstrate that the magnetic properties of the Fe/ZnSe(001) interface remain stable against thermal annealing up to 300 degrees C, a prerequisite for the future development of efficient spintronics devices.

Journal Article↗

PubMed2001

Protein-protein interactions of alpha-synuclein in brain homogenates and transfected cells.

alpha-Synuclein is a highly conserved presynaptic protein with probable roles in normal synaptic development and plasticity as well as neurodegenerative disease, although its molecular function is not yet clear. To identify potential protein binding partners of alpha-synuclein, we performed co-immunoprecipitations using a monoclonal antibody (H3C) against its C-terminus. More than 20 detectable proteins were specifically co-immunoprecipitated from zebra finch and mouse forebrain extracts. One of these, with relative mobility of 55 kDa, was identified through microsequencing as a mixture of alpha- and beta-tubulin. Tubulin was specifically recovered from a mouse forebrain cytosolic extract by a GST/alpha-synuclein fusion protein immobilized on glutathione-Sepharose beads. In the converse experiment, alpha-synuclein bound to a column prepared from purified bovine brain tubulin immobilized upon CNBr-Sepharose. alpha-Synuclein does not appear to bind assembled microtubules, however, as alpha-synuclein did not pellet with polymerized microtubules in a standard assay for microtubule-associated proteins. Likewise, when a fusion construct of alpha-synuclein and green fluorescent protein (GFP) was expressed in African green monkey kidney epithelial (CV-1) cells, the fusion protein did not colocalize with endogenous microtubules. We conclude that alpha-synuclein may interact specifically with heterodimeric tubulin, but not microtubules, in the neuronal cytosol.

Amino Acid Sequence↗

PubMed2001

Exposure to long chain polyunsaturated fatty acids triggers rapid multimerization of synucleins.

Detergent-stable multimers of alpha-synuclein have been found specifically in the brains of patients with Parkinson's disease and other neurodegenerative diseases. Here we show that recombinant alpha-synuclein forms multimers in vitro upon exposure to vesicles containing certain polyunsaturated fatty acid (PUFA) acyl groups, including arachidonoyl and docosahexaenoyl. This process occurs at physiological concentrations and much faster than in aqueous solution. PUFA-induced aggregation involves physical association with the vesicle surface via the large apolipoprotein-like lipid-binding domain that constitutes the majority of the protein. beta- and gamma-synucleins, as well as the Parkinson's disease-associated alpha-synuclein variants A30P and A53T, show similar tendencies to multimerize in the presence of PUFAs. Multimerization does not require the presence of any tyrosine residues in the sequence. The membrane-based interaction of the synucleins with specific long chain polyunsaturated phospholipids may be relevant to the protein family's physiological functions and may also contribute to the aggregation of alpha-synuclein observed in neurodegenerative disease.

Fatty Acids, Unsaturated↗

PubMed2001

When openness to experience and conscientiousness are related to creative behavior: an interactional approach.

This study adopted an interactional approach to understanding how 2 of the Five-Factor traits, openness to experience and conscientiousness, are related to creative behavior in the workplace. Openness to experience is theorized to result in high levels of creative behavior and conscientiousness is theorized to result in low levels of creative behavior when the situation allows for the manifestation of the trait influences. More specifically, the authors hypothesized that openness to experience would result in high levels of creative behavior if feedback valence were positive and job holders were presented with a heuristic task that allowed them to be creative. The authors also hypothesized that conscientiousness would result in low levels of creative behavior if supervisors engaged in close monitoring and coworkers were unsupportive. The authors tested their hypotheses in a sample of office workers, and 5 out of the 6 hypotheses were supported.

Adult↗

PubMed2000

Interaction of human alpha-Synuclein and Parkinson's disease variants with phospholipids. Structural analysis using site-directed mutagenesis.

alpha-Synuclein has been centrally implicated in neurodegenerative disease, and a normal function in developmental synaptic plasticity has been suggested by studies in songbirds. A variety of observations suggest the protein partitions between membrane and cytosol, a behavior apparently conferred by a conserved structural similarity to the exchangeable apolipoproteins. Here we show that the capacity to bind lipids is broadly distributed across exons 3, 4, and 5 (encoding residues 1-102). Binding to phosphatidylserine-containing vesicles requires the presence of all three exons, while binding to phosphatidic acid can be mediated by any one of the three. Consistent with a "class A2" helical binding mechanism, lipid association is disrupted by introduction of charged residues along the hydrophobic face of the predicted alpha-helix and also by biotinylation of conserved lysines (which line the interfacial region). Circular dichroism spectroscopy reveals a general correlation between the amount of lipid-induced alpha-helix content and the degree of binding to PS-containing vesicles. Two point mutations associated with Parkinson's disease have little (A30P) or no (A53T) effect on lipid binding or alpha-helicity. These results are consistent with the hypothesis that alpha-synuclein's normal functions depend on an ability to undergo a large conformational change in the presence of specific phospholipids.

Cloning, Molecular↗

PubMed1999

Synucleins in synaptic plasticity and neurodegenerative disorders.

Synucleins are small highly conserved proteins in vertebrates, especially abundant in neurons and typically enriched at presynaptic terminals. Three genes in humans produce closely related synuclein proteins, all of which share a large amphipathic domain capable of reversible binding to lipid vesicles. Alpha synuclein has been specifically implicated in neurodegenerative disease. Two point mutations are genetically linked to familial Parkinson's disease, and alpha synuclein appears to form the major fibrillary component of Lewy bodies. Alpha synuclein also contributes to the intracellular inclusions of multiple system atrophy, and a fragment has been found in senile plaques in Alzheimer's disease. Although their normal cellular functions are unknown, several observations suggest the synucleins may serve to integrate presynaptic signaling and membrane trafficking. Alpha synuclein has been identified as a potent and selective inhibitor of phospholipase D2, which produces phosphatidic acid (to which synuclein binds) and is believed to function in the partitioning of membranes between the cell surface and intracellular stores. We outline a hypothesis whereby synuclein supports localized, experience-dependent turnover of synaptic membranes. Such a process may be important for lifelong learning and memory functions and may be especially vulnerable to disruption in aging-associated neurodegenerative diseases.

Animals↗

PubMed1999

The reinforced laryngeal mask in paediatric outpatient dental surgery.

One hundred and twenty ASA I and II grade children aged 2-9 years scheduled for outpatient dental extractions under general anaesthesia were studied. They were allocated randomly to one of three groups for airway management: group R had anaesthesia with a reinforced laryngeal mask airway, group L with a standard laryngeal mask airway and group N with a nasal mask. Anaesthesia was induced in all children using halothane in 50% nitrous oxide with oxygen and maintained on halothane in 67% nitrous oxide with oxygen. An Ayre's T-piece with Jackson-Rees modification was used. The incidence of airway obstruction was significantly lower and surgical access significantly better with the reinforced laryngeal mask airway when compared with the standard laryngeal mask airway. However, the reinforced laryngeal mask airway was significantly more difficult to insert when compared with the standard laryngeal mask airway. On comparing the reinforced laryngeal mask airway with the nasal mask, there were significantly fewer episodes of airway obstruction, better oxygen saturation, less increase in heart rate and fewer arrhythmias in the reinforced laryngeal mask airway group. Total time for the procedures was the same for all three groups. Thus, the reinforced laryngeal mask airway was found to be a favourable alternative to the standard laryngeal mask airway and nasal mask for paediatric outpatient dental extractions.

Airway Obstruction↗

PubMed1998

Stabilization of alpha-synuclein secondary structure upon binding to synthetic membranes.

alpha-Synuclein is a highly conserved presynaptic protein of unknown function. A mutation in the protein has been causally linked to Parkinson's disease in humans, and the normal protein is an abundant component of the intraneuronal inclusions (Lewy bodies) characteristic of the disease. alpha-Synuclein is also the precursor to an intrinsic component of extracellular plaques in Alzheimer's disease. The alpha-synuclein sequence is largely composed of degenerate 11-residue repeats reminiscent of the amphipathic alpha-helical domains of the exchangeable apolipoproteins. We hypothesized that alpha-synuclein should associate with phospholipid bilayers and that this lipid association should stabilize an alpha-helical secondary structure in the protein. We report that alpha-synuclein binds to small unilamellar phospholipid vesicles containing acidic phospholipids, but not to vesicles with a net neutral charge. We further show that the protein associates preferentially with vesicles of smaller diameter (20-25 nm) as opposed to larger (approximately 125 nm) vesicles. Lipid binding is accompanied by an increase in alpha-helicity from 3% to approximately 80%. These observations are consistent with a role in vesicle function at the presynaptic terminal.

Amino Acid Sequence↗

PubMed1998

The synucleins: a family of proteins involved in synaptic function, plasticity, neurodegeneration and disease.

Synuclein proteins are produced, in vertebrates, by three genes. They share structural resemblance to apolipoproteins, but are abundant in the neuronal cytosol and present in enriched amounts at presynaptic terminals. Synucleins have been specifically implicated in three diseases:Alzheimer's (AD), Parkinson's (PD) and breast cancer. In AD, a peptide derived from alpha-synuclein forms an intrinsic component of plaque amyloid. In PD, an alpha-synuclein allele is genetically linked to several independent familial cases, and the protein appears to accumulate in Lewy bodies. In breast cancer, increased expression of gamma-synuclein correlates with disease progression. In songbirds, alpha-synuclein expression is correlated with plasticity in the developing song control system. Although the normal function of synucleins is unknown, a role in membrane plasticity seems likely.

Animals↗

PubMed1998

Nigral and cortical Lewy bodies and dystrophic nigral neurites in Parkinson's disease and cortical Lewy body disease contain alpha-synuclein immunoreactivity.

A mutation in the alpha-synuclein gene has recently been linked to some cases of familial Parkinson's disease (PD). We characterized the expression of this presynaptic protein in the midbrain, striatum, and temporal cortex of control, PD, and dementia with Lewy bodies (DLB) brain. Control brain showed punctate pericellular immunostaining. PD brain demonstrated alpha-synuclein immunoreactivity in nigral Lewy bodies, pale bodies and abnormal neurites. Rare neuronal soma in PD brain were immunoreactive for alpha-synuclein. DLB cases demonstrated these findings as well as alpha-synuclein immunoreactivity in cortical Lewy bodies and CA2-3 neurites. These results suggest that, even in sporadic cases, there is an early and direct role for alpha-synuclein in the pathogenesis of PD and the neuropathologically related disorder DLB.

Aged↗

PubMed1997

An extracellular matrix response element in the promoter of the LpS1 genes of the sea urchin Lytechinus pictus.

The extracellular matrix (ECM) has been shown to play an important role in development and tissue-specific gene expression, yet the mechanism by which genes receive signals from the ECM is poorly understood. The aboral ectoderm-specific LpS1-alpha and -beta genes of Lytechinus pictus , members of the Spec gene family, provide an excellent model system to study ECM- mediated gene regulation. Disruption of the ECM by preventing collagen deposition using the lathrytic agent beta-aminopropionitrile (BAPN) inhibits LpS1 gene transcription. LpS1 transcription resumes after removal of BAPN and subsequent collagen reformation. Using a chloramphenicol acetyltransferase (CAT) reporter gene assay, we show that a 125 bp region of the LpS1-beta promoter from -108 to +17 contains an ECM response element (ECM RE). Insertion of the 125 bp region into the promoter of the metallothionein gene of L. pictus, a gene unaffected by ECM disruption, caused the fused promoter to become ECM dependent. As with the endogenous LpS1 genes, CAT activity directed by the fused LpS1-beta promoter resumed in embryos recovered from ECM disruption. A mutation in a cis -acting element called the proximal G-string, which lies in the 125 bp region, caused CAT activity levels in ECM-disrupted embryos to equal that of the wild-type LpS1-bet apromoter in ECM-intact embryos. These results suggest that the intact ECM normally transmits signals to inhibit repressor activity at the proximal G-string in aboral ectoderm cells. Consistent with these results were our findings which showed that in addition to expression in the aboral ectoderm, the proximal G-string mutation caused expression of the CAT gene in oral ectoderm cells. These studies suggested that the proximal G-string serves as a binding site for negative regulation of the LpS1 genes in oral ectoderm during development. We also examined trans -acting factors binding the proximal G-string following ECM disruption. Band shift gels revealed a predominant set of slower migrating nuclear proteins from ECM-disrupted embryos which bound the proximal G-string. This work suggested that ECM disruption initiates signaling that induces a repressor to bind the ECM RE and/or modifies ECM RE binding proteins, which in turn represses LpS1 gene activity.

Animals↗

PubMed1997

Delayed localization of synelfin (synuclein, NACP) to presynaptic terminals in cultured rat hippocampal neurons.

Synelfin is a presynaptic protein of unknown function that is differentially regulated in the avian song control circuit during the critical period for song learning; in humans, it gives rise to an amyloidogenic peptide found in senile plaques of Alzheimer's disease. To gain insight into the potential involvement of synelfin in synapse development, we investigated its expression in neurons cultured from the embryonic rat hippocampus. These neurons express a variety of defined synaptic proteins, and form numerous synaptic connections after several days in culture. Synapsin I, a synaptic vesicle-associated protein, was detected within one day after the neurons were put in culture, but significant immunoreactivity for synelfin was not detected until approximately 5 days in vitro (DIV). By 3 DIV, synapsin-positive puncta (previously shown to correspond to presynaptic specializations) were detected surrounding the soma and proximal dendritic processes, whereas comparable aggregations of synelfin did not appear until several days later. By 14 DIV the punctate concentrations of synelfin and synapsin overlapped completely. Thus synelfin is expressed in these cultured neurons and eventually becomes localized to presynaptic terminals, but it is absent from these specializations when they first form. We conclude that presynaptic terminals can change in molecular composition, and that synelfin is associated with later stages in synaptic development or modulation.

Amyloid↗

PubMed1996

USF in the Lytechinus sea urchin embryo may act as a transcriptional repressor in non-aboral ectoderm cells for the cell lineage-specific expression of the LpS1 genes.

Expression of the aboral ectoderm-specific LpS1 gene in Lytechinus was used to study lineage-specific transcriptional regulation during sea urchin development. Band shift assays using anti-USF antibody showed that a USF-like protein bound the USF core sequence 5'-CACGTG-3' in the promoter of the LpS1 gene. DNA constructs consisting of a wild-type LpS1 promoter and the same LpS1 promoter with a mutated USF binding site fused to the bacterial chloramphenicol acetyltransferase reporter gene were tested. The mutation in the USF binding site caused an increase in chloramphenicol acetyltransferse activity. We selected a clone that encodes USF, LvUSF, from a gastrula-stage cDNA library representing Lytechinus variegatus. Transactivation experiments, in which LvUSF RNA or a DNA construct consisting of the LvUSF cDNA clone fused to the Lytechinus pictus metallothionein promoter coinjected with the wild-type or mutated LpS1 promoter-chloramphenicol acetyltransferase gene construct, showed that chloramphenicol acetyltransferase activity from the wild-type construct was repressed, while the construct mutated at the USF binding site was active. The same wild-type and mutated LpS1 promoter DNA fragments ligated to the green fluorescent protein reporter gene were used to examine spatial expression. The reporter gene constructs containing the mutated USF binding site were expressed inappropriately in all cell types including the gut and oral ectoderm in gastrula and larva stage embryos, while the wild-type constructs were expressed primarily in the aboral ectoderm. USF was expressed in all cells of the early embryo and in all tissues except the aboral ectoderm in later embryos. The data are consistent with a model depicting Lytechinus USF, as a temporal and spatial regulator by repressing LpS1 gene transcription in non-aboral ectoderm cells.

Amino Acid Sequence↗

PubMed1996

Two distinct forms of USF in the Lytechinus sea urchin embryo do not play a role in LpS1 gene inactivation upon disruption of the extracellular matrix.

Recent studies in our laboratory indicated that the upstream stimulatory factor (USF) in the sea urchin embryo of Lytechinus acts as a transcriptional repressor for the aboral ectoderm-specific expression of the LpS1 genes. Disruption of the extracellular matrix (ECM) arrests development prior to gastrulation and inactivates the LpS1 genes. We wanted to determine whether the inactivation of the LpS1 genes by ECM disruption may be due to an increase in USF expression. In the course of the investigation, a second L, variegatus USF cDNA clone (LvUSF2) was isolated and sequenced. The deduced amino acid sequence of LvUSF2 is nearly identical to LvUSF1 except at the amino end, where they are sharply divergent. Like LvUSF1, LvUSF2 has a USF-specific, a basic/hefixloop-helix, and a leucine zipper domain. Genomic DNA blots indicated that the two cDNA clones are derived from one gene, which suggested that the Lytechinus USF1 and USF2 mRNAs, of approximately 6.0 and 4.0 kb, respectively, are the result of differential RNA splicing. ECM disruption in Lytechinus embryos caused a relative drop in USF RNA accumulation levels to approximately 60% of control embryos, while LpS1 RNA accumulation levels dropped to less than 5%. USF protein levels and DNA binding activities in ECM-disrupted embryos also dropped to approximately 60% to that of control embryos. A mutation at the USF binding site in an LpS1 promoter-chloramphenicol acetyl transferase (CAT) fusion DNA construct did not cause a relative increase in CAT activity in ECM disrupted embryos. These results suggest that the induced drop in LpS1 gene expression by ECM disruption is not due to an increase in the repressive activity of USF.

Amino Acid Sequence↗

PubMed1996

The experience of work and turnover intentions: interactive effects of value attainment, job satisfaction, and positive mood.

The researchers proposed that the phenomenological experience of work is multifaceted and that simultaneously considering the combined effects of the different aspects of the work experience holds considerable promise for increasing understanding of turnover intentions and other organizationally relevant outcomes. Research in subjective well-being and other areas suggests that there are 3 important aspects of the experience of work: attainment of values, attitudes, and moods. It was hypothesized and found that the relationship between job satisfaction and turnover intentions is jointly moderated by value attainment and positive mood. The nature of this triple interaction was such that the job satisfaction-turnover intention relationship was strongest when workers' jobs did not help them to attain terminal values and positive moods were experienced, and the relationship was weakest when jobs helped workers to attain terminal values and positive moods were experienced.

Adult↗

PubMed1996

Characterization of the precursor protein of the non-A beta component of senile plaques (NACP) in the human central nervous system.

A novel and highly conserved presynaptic protein has been independently described in rodents (synuclein/SYN-1), songbirds (synelfin), and humans (the precursor protein of the non-A beta component of senile plaques, NACP); a fragment of the latter has been detected in senile plaques in Alzheimer's disease (AD). We characterized the expression of NACP in human AD and non-AD brain. A subcellular fractionation study demonstrated that NACP was mainly localized to cytosolic fractions of human temporal cortex. NACP was also detectable in various membrane and vesicular fractions, suggesting that the protein was associated with membrane structures including synaptic vesicles. Pericellular immunostaining of the neuropil was observed in neocortical and limbic regions, supporting a synaptic localization. Senile plaques in AD brains were not immunoreactive, and confocal microscopy suggested a loss of NACP immunoreactivity in cored plaques. No difference was found in the amount of protein in AD and control frontal cortex, as measured by immunoblotting. PCR analysis showed that the full-length mRNA product was the major splice form in both AD and control human brains. Thus, despite the association of a hydrophobic fragment of NACP with senile plaques, our data suggest that the precursor itself is not a significant component of plaques and NACP synthesis is not substantially altered in AD. Nevertheless, the protein is an abundant component of synaptic regions prone to degeneration in AD, and may have a role in the expression or advancement of the disease.

Adult↗

PubMed1995

An ECM-bound, PDGF-like growth factor and a TGF-alpha-like growth factor are required for gastrulation and spiculogenesis in the Lytechinus embryo.

Growth factors and the extracellular matrix have been shown to fulfill vital developmental roles in many embryonic systems. Our hypothesis is that a developmental role played by the extracellular matrix in sea urchins may be the binding of a PDGF-like growth factor to promote signaling activity. We report here that anti-human PDGF-B antibodies and anti-human TGF-alpha antibodies immunoprecipitated specific proteins isolated from Lytechinus embryos. Addition of these antibodies to Lytechinus embryos inhibited gastrulation and spiculogenesis. The embryos are sensitive to the antibodies from the four-cell through the hatching blastula stages, which suggests that the TGF-alpha-like and PDGF-like ligands are required for the early differentiation of the gut and spicules. We present evidence that the PDGF-like growth factor depends on the extracellular matrix for signaling activity. Synthetic peptides representing the heparan sulfate proteoglycan binding sequence on human PDGF-B were added to Lytechinus embryo cultures to compete for binding sites with the endogenous PDGF-like growth factor. The experimental peptide inhibited gastrulation and caused radially arranged multiple spicules to form. Development was unaffected by a control peptide. These studies support our hypothesis and suggest that TGF-alpha-like and PDGF-like growth factors induce signaling events required for sea urchin gastrulation and spiculogenesis and suggest that an extracellular matrix-associated PDGF-like growth factor is involved in differentiation along the oral-aboral axis.

Amino Acid Sequence↗