Figure 4.

A region in the 3′ UTR of TDP-43 mediates autoregulation (A) a schematic diagram of the GFP-3′ UTR wt and Δ369 and Δ669 constructs. Wild-type TDP-43 coding sequences are shown in grey. The lower panel shows the splicing events that lead to the Δ1812 mRNA and protein isoforms. (B) GFP protein expression when the GFP-3′ UTR wt and Δ369 and Δ669 constructs were transfected into the HEK293 stable cell lines following induction (+) of the TG-TDP43 transgene. Transfection efficiencies were normalized by co-transfecting a GFP expression vector (lower panel). In the GFP-3′ UTR Δ369 and Δ669 (−) and (+) lanes, the western blot also contains an additional 40 kDa protein band that was called Δ1812. The different proteins were detected with anti-GFP (C) shows a northern blot analysis of the transcripts derived from the GFP-3′ UTR wt, Δ369, and Δ669 constructs in the HEK293 stable cell lines before (−) and after Tet induction (+). All constructs produce two major mRNA species because of the usage of different polyadenylation sites. In addition to these species, the GFP-3′ UTR Δ369 and Δ669 constructs also produce an aberrantly spliced mRNA isoform (Δ1812) that accounts for the production of the aberrant protein present in the western blots (B).