Regulation of DYT1 gene expression by the Ets family of transcription factors

Abstract

The DYT1 gene encodes for torsinA, a protein with widespread tissue distribution, involved in early onset dystonia (EOD). Numerous studies have focused on torsinA function but no information is available on its transcriptional regulation. We cloned mouse and human 5′‐upstream DYT1 DNA fragments, exhibiting high transcriptional activity, as well as tissue specificity. We identified a proximal minimal DYT1 promoter within −141 bp for mouse and −191 bp for human with respect to the ATG codon. Primer extension analysis indicated multiple transcription start sites. In silico analysis of approximately 500 bp 5′‐upstream DYT1 fragment demonstrated lack of a classical TATA or CAAT box and the presence of a highly conserved direct repeat of two Ets binding cores within −86 bp to −77 bp and −78 bp to −69 bp of the mouse and human DYT1 gene, respectively. A single or a two base nucleotide alteration within the downstream Ets core resulted in approximately 90% (mouse) or 45–60% (human) drop in activity. Interestingly, a 3‐bp distance increase between the two Ets cores dramatically decreased transcriptional activity which was partially restored when the distance was increased up to 10 bp. Ets‐like dominant negatives confirmed the Ets factors as DYT1 transcriptional activators.

Localization of immunoreactive, dystonia-associated protein torsinA near the Golgi apparatus of cultured rodent astrocytes

“…1B), with a minor component surrounding the nuclei (perinuclear). This finding is consistent with results from an earlier study in cultured human astrocytes (Armata et al, 2008).…”

“…Nevertheless, the mis-localization model of ΔE-torsinA has not been tested in astrocytes of mammalian brains. Human WT-torsinA was overexpressed in human astrocytes in primary culture, and was found to be distributed diffusely in the cytoplasm and co-localized with an astrocyte marker, the cytoskeletal, intermediate-filament protein glial fibrillary acidic protein (GFAP), in both the cell body and the processes (Armata et al, 2008). While this result is consistent with the assumed localization of WT-torsinA to the ER, ΔE-torsinA has not been overexpressed in astrocytes.…”

“…Since overexpressed WT-torsinA is co-localized with GFAP in human astrocytes (Armata et al, 2008), we also double-stained the cells for endogenous torsinA and GFAP (Fig. 5B).…”

“…With respect to astrocytes, the outcome of torsinA overexpression had been reported in only one study. Human WT-torsinA had been overexpressed in cultured human astrocytes of a non-specified brain region (Armata et al, 2008). The signal was diffuse throughout the cytoplasm and co-localized with GFAP.…”

Molecular pathways in dystonia

“…The human TOR1A promoter has been characterized (Armata et al, 2008) and recently shown to contain two potential binding sites for THAP1: an inverted bipartite motif (-111 bp to -101 bp from the putative transcriptional start site) and a more upstream, nonconserved motif (-259 bp to -252 bp from the start site; Gavarini et al, 2010; Kaiser et al, 2010). Both studies demonstrated that wild-type THAP1 binds these sequences, whereas multiple DYT6 mutant forms of THAP1 do not.…”