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Functional Genomics of Human Bronchial Epithelial Cells Directly Interacting with Conidia of Aspergillus Fumigatus

Peer reviewed: 
Yes, item is peer reviewed.
Date created: 
2010
Abstract: 

Background: Aspergillus fumigatus (A. fumigatus) is a ubiquitous fungus which reproduces asexually by releasingabundant airborne conidia (spores), which are easily respirable. In allergic and immunocompromised individuals A.fumigatus can cause a wide spectrum of diseases, including allergic bronchopulmonary aspergillosis, aspergilloma andinvasive aspergillosis. Previous studies have demonstrated that A. fumigatus conidia are internalized by macrophagesand lung epithelial cells; however the exact transcriptional responses of airway epithelial cells to conidia are currentlyunknown. Thus, the aim of this study was to determine the transcriptomic response of the human bronchial epithelialcell line (16HBE14o-) following interaction with A. fumigatus conidia. We used fluorescence-activated cell sorting (FACS)to separate 16HBE14o- cells having bound and/or internalized A. fumigatus conidia expressing green fluorescentprotein from cells without spores. Total RNA was then isolated and the transcriptome of 16HBE14o- cells was evaluatedusing Agilent Whole Human Genome microarrays.Results: Immunofluorescent staining and nystatin protection assays demonstrated that 16HBE14o- cells internalized30-50% of bound conidia within six hrs of co-incubation. After FAC-sorting of the same cell culture to separate cellsassociated with conidia from those without conidia, genome-wide analysis revealed a set of 889 genes showingdifferential expression in cells with conidia. Specifically, these 16HBE14o- cells had increased levels of transcripts fromgenes associated with repair and inflammatory processes (e.g., matrix metalloproteinases, chemokines, andglutathione S-transferase). In addition, the differentially expressed genes were significantly enriched for Gene Ontologyterms including: chromatin assembly, G-protein-coupled receptor binding, chemokine activity, and glutathionemetabolic process (up-regulated); cell cycle phase, mitosis, and intracellular organelle (down-regulated).Conclusions: We demonstrate a methodology using FACs for analyzing the transcriptome of infected and uninfectedcells from the same cell population that will provide a framework for future characterization of the specific interactionsbetween pathogens such as A. fumigatus with human cells derived from individuals with or without underlying diseasesusceptibility.

Document type: 
Article

A Glycine Zipper Motif Mediates the Formation of Toxic Beta-Amyloid Oligomers in Vitro and in Vivo

Peer reviewed: 
Yes, item is peer reviewed.
Date created: 
2011
Abstract: 

Background: The b-amyloid peptide (Ab) contains a Gly-XXX-Gly-XXX-Gly motif in its C-terminal region that hasbeen proposed to form a “glycine zipper” that drives the formation of toxic Ab oligomers. We have tested thishypothesis by examining the toxicity of Ab variants containing substitutions in this motif using a neuronal cell line,primary neurons, and a transgenic C. elegans model.Results: We found that a Gly37Leu substitution dramatically reduced Ab toxicity in all models tested, as measuredby cell dysfunction, cell death, synaptic alteration, or tau phosphorylation. We also demonstrated in multiplemodels that Ab Gly37Leu is actually anti-toxic, thereby supporting the hypothesis that interference with glycinezipper formation blocks assembly of toxic Ab oligomers. To test this model rigorously, we engineered second sitesubstitutions in Ab predicted by the glycine zipper model to compensate for the Gly37Leu substitution andexpressed these in C. elegans. We show that these second site substitutions restore in vivo Abtoxicity, furthersupporting the glycine zipper model.Conclusions: Our structure/function studies support the view that the glycine zipper motif present in the Cterminalportion of Ab plays an important role in the formation of toxic Ab oligomers. Compounds designed tointerfere specifically with formation of the glycine zipper could have therapeutic potential.

Document type: 
Article

Association Testing Of Copy Number Variants in Schizophrenia and Autism Spectrum Disorders

Peer reviewed: 
Yes, item is peer reviewed.
Date created: 
2012
Abstract: 

Background: Autism spectrum disorders and schizophrenia have been associated with an overlapping set of copynumber variant loci, but the nature and degree of overlap in copy number variants (deletions compared toduplications) between these two disorders remains unclear.Methods: We systematically evaluated three lines of evidence: (1) the statistical bases for associations of autismspectrum disorders and schizophrenia with a set of the primary CNVs thus far investigated, from previous studies;(2) data from case series studies on the occurrence of these CNVs in autism spectrum disorders, especially amongchildren, and (3) data on the extent to which the CNVs were associated with intellectual disability anddevelopmental, speech, or language delays. We also conducted new analyses of existing data on these CNVs inautism by pooling data from seven case control studies.Results: Four of the CNVs considered, dup 1q21.1, dup 15q11-q13, del 16p11.2, and dup 22q11.21, showed clearstatistical evidence as autism risk factors, whereas eight CNVs, del 1q21.1, del 3q29, del 15q11.2, del 15q13.3, dup16p11.2, dup 16p13.1, del 17p12, and del 22q11.21, were strongly statistically supported as risk factors forschizophrenia. Three of the CNVs, dup 1q21.1, dup 16p11.2, and dup 16p13.1, exhibited statistical support as riskfactors for both autism and schizophrenia, although for each of these CNVs statistical significance was nominal fortests involving one of the two disorders. For the CNVs that were statistically associated with schizophrenia but werenot statistically associated with autism, a notable number of children with the CNVs have been diagnosed withautism or ASD; children with these CNVs also demonstrate a high incidence of intellectual disability anddevelopmental, speech, or language delays.Conclusions: These findings suggest that although CNV loci notably overlap between autism and schizophrenia,the degree of strongly statistically supported overlap in specific CNVs at these loci remains limited. These analysesalso suggest that relatively severe premorbidity to CNV-associated schizophrenia in children may sometimes bediagnosed as autism spectrum disorder.

Document type: 
Article