cDNA clones corresponding to the polypeptide that has been shown to be an endogenous diazepam binding inhibitor and may act as a physiological ligand for the benzodiazepine/beta-carboline receptor have been isolated from bacteriophage lambda recombinant libraries from rat hypothalamus, total brain, and liver. The clones contain an open reading frame corresponding to 87 amino acids. A signal sequence is not present. In addition to high levels of mRNA in various brain regions, RNA blot analysis reveals an abundance of diazepam binding inhibitor mRNA in many peripheral organs (e.g., testes, kidney, liver, and heart) that are known to be rich in peripheral benzodiazepine recognition sites. The size of the mRNA in all tissue examined is approximately 0.7 kilobase. Southern blot analysis of genomic DNA suggests the presence of about six genes in the rat, some of which may be pseudogenes.

This study provides proof-of-concept that whole-blood profiling can generate an AD-associated classification signature via the specific relative expression of biologically relevant RNAs. Such a signature will need to be validated with extended patient cohorts, and evaluated to learn whether it can differentiate AD from others types of dementia.

G protein-coupled receptors (GPCRs) are the largest signaling family in the genome, serve an expansive array of functions, and are targets for Ϸ50% of current therapeutics. In many tissues, such as airway smooth muscle (ASM), complex, unexpected, or paradoxical responses to agonists/antagonists occur without known mechanisms. We hypothesized that ASM express many more GPCRs than predicted, and that these undergo substantial alternative splicing, creating a highly diversified receptor milieu. Transcript arrays were designed detecting 434 GPCRs and their predicted splice variants. In this cell type, 353 GPCRs were detected (including 111 orphans), with expression levels varying by Ϸ900-fold. Receptors used for treating airway disease were expressed lower than others with similar signaling properties, indicating potentially more effective targets. A disproportionate number of Class-A peptide-group receptors, and those coupling to G q/11 or Gs (vs. Gi), was found. Importantly, 192 GPCRs had, on average, five different expressed receptor isoforms because of splicing events, including alternative splice donors and acceptors, novel introns, intron retentions, exon(s) skips, and novel exons, with the latter two events being most prevalent. The consequences of splicing were further investigated with the leukotriene B4 receptor, known for its aberrant responsiveness in lung. We found transcript expression of three variants because of alternative donor and acceptor splice sites, representing in-frame deletions of 38 and 100 aa, with protein expression of all three isoforms. Thus, alternative splicing, subject to conditional, temporal, and cell-type regulation, is a major mechanism that diversifies the GPCR superfamily, creating local recepteromes with specialized environments.pharmacogenetics ͉ signal transduction ͉ splice variants

Biomarkers have gained an increased importance in the past years in helping physicians to diagnose Alzheimer's disease (AD). This study was designed to identify a blood-based, transcriptomic signature that can differentiate AD patients from control subjects. The performance of the signature was then evaluated for robustness in an independent blinded sample population. RNA was extracted from 177 blood samples (90 AD patients and 87 controls) and gene expression profiles were generated using the human Genome-Wide Splice Array™. These profiles were used to establish a signature to differentiate AD patients from controls. Subsequently, prediction results were optimized by establishing grey zone boundaries that discount prediction scores near the disease status threshold. Signature validation was then performed on a blinded independent cohort of 209 individuals (111 AD and 98 controls). The AclarusDx™ signature consists of 170 probesets which map to 136 annotated genes, a significant number of which are associated with inflammatory, gene expression, and cell death pathways. Additional signature genes are known to interact with pathways involved in amyloid and tau metabolism. The validation sample set, after removal of 45 individuals with prediction profile scores within the grey zone, consisted of 164 subjects. The AclarusDx™ performance on this validation cohort had a sensitivity of 81.3% (95% CI: [73.3%; 89.3%]); and a specificity of 67.1% (95% CI: [56.3%; 77.9%]). AclarusDx™ is a non-invasive blood-based transcriptomic test that, in combination with standard assessments, can provide physicians with objective information to support the diagnosis of AD.

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