Abstract
Tar DNA Binding Protein-43 (TDP-43) is a principle component of inclusions in many cases of frontotemporal lobar degeneration (FTLD-U) and amyotrophic lateral sclerosis (ALS). TDP-43 resides predominantly in the nucleus, but in affected areas of ALS and FTLD-U central nervous system, TDP-43 is aberrantly processed and forms cytoplasmic inclusions. The mechanisms governing TDP-43 inclusion formation are poorly understood. Increasing evidence indicates that TDP-43 regulates mRNA metabolism by interacting with mRNA binding proteins that are known to associate with RNA granules. Here we show that TDP-43 can be induced to form inclusions in cell culture and that most TDP-43 inclusions co-localize with SGs. SGs are cytoplasmic RNA granules that consist of mixed protein - RNA complexes. Under stressful conditions SGs are generated by the reversible aggregation of prion-like proteins, such as TIA-1, to regulate mRNA metabolism and protein translation. We also show that disease-linked mutations in TDP-43 increased TDP-43 inclusion formation in response to stressful stimuli. Biochemical studies demonstrated that the increased TDP-43 inclusion formation is associated with accumulation of TDP-43 detergent insoluble complexes. TDP-43 associates with SG by interacting with SG proteins, such as TIA-1, via direct protein-protein interactions, as well as RNA-dependent interactions. The signaling pathway that regulates SGs formation also modulates TDP-43 inclusion formation. We observed that inclusion formation mediated by WT or mutant TDP-43 can be suppressed by treatment with translational inhibitors that suppress or reverse SG formation. Finally, using Sudan black to quench endogenous autofluorescence, we also demonstrate that TDP-43 positive-inclusions in pathological CNS tissue co-localize with multiple protein markers of stress granules, including TIA-1 and eIF3. These data provide support for accumulating evidence that TDP-43 participates in the SG pathway.
Highlights
Tar DNA Binding Protein-43 (TDP-43) is the principle protein component of inclusions in amyotrophic lateral sclerosis (ALS) and ubiquitin positive frontotemporal lobar degeneration (FTLDU) [1]
To test whether inclusions would form in neuronal cells, human BE-M17 neuroblastoma cells were transfected with with GFP or TDP-43 (WT) TDP-43, TDP-4386–414 or TDP-43216–414 constructs N-terminally tagged with EGFP (Fig 1)
Full length WT TDP-43 predominently localized to the nucleus under basal conditions with only 10% of the cells exhibiting TDP-43 inclusions (Fig. 2A; Supplemental figure S1 shows an example of a cell with some cytoplasmic TDP-43 inclusions)
Summary
TDP-43 is the principle protein component of inclusions in ALS and ubiquitin positive frontotemporal lobar degeneration (FTLDU) [1]. TDP-43 is a 414 amino acid nuclear protein encoded by the TARDBP gene on chromosome 1. It is ubiquitously expressed in all tissues, it highly expresses in the brain and kidney [4]. TDP-43 is an mRNA binding protein that plays important functions in regulating mRNA metabolism involved in several functions, including transcriptional repression, exon skipping and RNA splicing [5,6]. It contains two RNA binding domains and a glycine rich domain at the C terminus. The mislocalization of TDP-43 in the cytoplasm highlights important gaps in our knowledge of TDP-43 biology
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
