Single molecule profiling of tau gene expression in Alzheimer’s disease

Abstract

Tau is a microtubule-associated protein that is important for establishing and maintaining neuronal morphology. In addition to its role in normal cells, tau protein is involved in many neurodegenerative diseases, e.g. Alzheimer's disease (AD) and frontotemporal dementia, as the main component of intraneuronal aggregates. Alternative splicing of tau gene in the brain can give rise to at least six protein variants. A causative role of skewed tau exon 10 inclusion has been defined in frontotemporal dementia; however, no link was established between the aberrant splicing of tau and AD. Here, we applied a single-molecule-based technology, polymerase colony or polony, to simultaneously monitor tau splicing variant and haplotype profile in sporadic AD and normal brains. We found that the coordinated expression of tau exons 2 and 10 is altered in AD. Additional investigations of cis and trans mechanisms of this observation revealed a decreased protein expression of a known tau splicing factor, htra2-beta-1 in AD, thereby implicating a trans mechanism. Our results demonstrate that dysregulation of combinatorial splicing might serve as a signature for aging-related diseases, and the polony assay could be widely adapted for the study of other tauopathies. Furthermore, splicing-based therapeutics is an emerging area of drug development, and a well-defined and quantitative assay for monitoring single-gene transcriptome will be relevant for such development. Alternative pre-mRNA splicing is a major contributor of mammalian proteome diversity. Combinatorial usage of multiple alternatively spliced exons of a transcript can lead to exponential accumulation of protein variants, which may play distinct roles in establishing and maintaining cellular identity (Lander et al. 2001). One such locus encodes for the microtubule-associated protein, tau. In the human CNS, at least six tau isoforms can be produced as the result of combinatorial inclusion of its exons 2, 3, and 10 (Fig. 1a). Although the function of individual tau variants remains to be fully determined, dysregulation of tau splicing is often observed in neurodegenerative diseases with aberrant tau deposition, including frontotemporal dementia (FTD), Pick's disease, and progressive supranuclear palsy (Brandt et al. 2005). In FTD, several mutations in the tau gene have been shown to not only promote tau aggregation but also skew tau splicing (D'Souza and Schellenberg 2005), prompting the investigation of tau splicing in other tau pathology-related disorders like Alzheimer's disease (AD).Alternative splicing of the tau gene in AD patients had been previously examined using in situ hybridization (Goedert et al. 1989a,b), microarray analysis (Ginsberg et al. Yasojima et al. 1999;Boutajangout et al. 2004;Umeda et al. 2004;Connell et al. 2005;Glatz et al. 2006;Ingelsson et al. 2006); most of these studies did not detect differences in AD, although several experiments suggested an increase in exon 10 in tangle affected neurons or brain areas (Yasojima et al. 1999;Glatz et a...

RNA Helicase p68 (DDX5) Regulates tau Exon 10 Splicing by Modulating a Stem-Loop Structure at the 5′ Splice Site

“…More than 30 mutations have been identified among FTDP-17 patients, many of which affect tau exon 10 splicing (26,45,54,60,61,91,92,97; reviewed in references 2, 53, and 66). In addition, a number of studies have reported an altered Tau4R-to-Tau3R ratio or aberrant tau exon 10 splicing in a range of sporadic tauopathies, including progressive supranuclear palsy (PSP), corticobasal degeneration (CBD), multiple system tauopathy with dementia (MSTD), and argyrophilic grain disease (AGD), and even in some Alzheimer's patients, among others (20,39,101; reviewed in references 2, 12, 36, and 78 and references therein). It is interesting that a significant fraction of these patients show increased levels of Tau4R without detectable mutations in the tau gene.…”

Glycogen synthase kinase‐3β and tau genes interact in Alzheimer's disease

“…Another possibility is that by using brain samples from patients with tauopathies, one cannot differentiate between cis-acting elements and trans-acting splicing factors that increase splicing in of MAPT exon 10. 30,31 From the known effect on gene expressions by the GSK3B 19 and MAPT haplotypes (see Fig 1A), we propose the following model for the epistatic interaction between the two genes as shown in Figure 3. We propose that the downstream effect of discordant GSK-3␤ and Tau levels is altered interaction with other GSK-3␤ substrates, where excess Tau is able to sequester GSK-3␤ and shunt it toward cellular pools that will have functional effects on other substrates of GSK-3␤.…”

Single-Cell and Regional Gene Expression Analysis in Alzheimer’s Disease

“…In the human central nervous system (CNS), there are at least six isoforms of tau that are classified as either 3R (containing three tandem repeats) or 4R (containing four tandem repeats) (Goedert et al 1989). Conrad et al (2007) used a polymerase colony (polony)-based exon profiling platform, which allows the quantitative measure of combinatorial splicing of tau isoforms at a single molecule level. The assay uses an acrylamide gel matrix with randomly arrayed DNA templates that is processed via in-gel PCR amplification and is followed by an exon-specific oligonucleotide hybridization/single base extension.…”