TAR DNA-binding protein 43

TDP-43 is 414 amino acid residues long. It consists of four domains: an N-terminal domain spanning residues 1–76 (NTD) with a well-defined fold that has been shown to form a dimer or oligomer; two highly conserved folded RNA recognition motifs spanning residues 106–176 (RRM1) and 191–259 (RRM2), respectively, required to bind target RNA and DNA; an unstructured C-terminal domain encompassing residues 274–414 (CTD), which contains a glycine-rich region, is involved in protein-protein interactions, and harbors most of the mutations associated with familial amyotrophic lateral sclerosis. The entire protein devoid of large solubilising tags has been purified. The full-length protein is a dimer. but shortly after it was argued that in the context of FTLD-TDP the protein involved could be TMEM106B (which has been also resolved with cryo-EM), rather than of TDP-43. N-Terminal domain (NTD) The NTD located between residues 1 and 76 is involved in TDP-43 polymerization. Indeed, dimers are formed by head-to-head interactions between NTDs, and the polymer thus obtained allows for pre-mRNA splicing. However, further oligomerization brings to more toxic accumulates. This process of polymerization into dimers, larger forms or just stabilizing monomers is dependent on TDP-43 conformational equilibrium between monomers, homodimers and oligomers. Hence, in TDP-43 diseased cells, TDP-43's over-expression leads to the NTD showing high propensity to aggregate. Contrary to this, in normal cells, normal levels of TDP-43 allow for folded NTD, preventing aggregates and polymers formation. More recently, this domain was found to have a ubiquitin-like structure. It bears 27,6% of homology with Ubiquitin-1 and a β1-β2-α1-β3-β4-β5-β6 + 2*SO42- form. Ubiquitin-like domain are usually associated with a greater affinity for RNA/DNA. However, in the unique case of TDP-43, the Ubiquitin-like NTD binds directly to ssDNA. This interaction permits the conformational equilibrium cited higher to shift towards non-aggregated forms. The domain spanning from [1,80] has a solenoid-like structure which sterically impedes interactions between aggregation prone C-term regions. Mitochondrial localization signal There are six mitochondrial localization signals to be accounted on TDP-43's amino acid sequence, although only M1, M3, and M5 were shown to be essential for mitochondrial localization. Indeed, their ablation leads to a lessened mitochondrial localization. These localizing sequences are found on the following amino acids: M1: [35, 41], M2: [105, 112], M3: [146-150], M4: [228, 235], M5: [294, 300], M6: [228, 236]. Nuclear localization signal (NLS) The nuclear localization signal (NLS) domain is located between residues 82 and 98 is of critical importance in ALS, and such is witnessed by the depletion or the mutations (notably A90V) of this domain, which cause loss-of-function from nucleus and promote aggregating, two processes very likely to conduct to TDP-43's toxic gain of function. Nuclear export signal (NES) The nuclear export signal is located between residues 239 and 251 sequence probably bears a role in TDP-43's shuttling function, and was recently found using a prediction algorithm. Disordered glycin rich C-terminal domain (CTD) The Disordered Glycin Rich C-terminal domain is located between residues 277 and 414. Much like 70 other RNA binding proteins, TDP-43 bears a Q/N rich domain [344, 366] which resembles yeast prion sequence. This sequence is called a Prion-Like Domain (PLD). PLDs are low complexity sequences that have been reported to mediate gene regulation via Liquid-Liquid Phase Transition (LLP) thus driving RNP granule assembly. It is here noted that LLP are reversible phenomenons of de-mixing a solution into two distinct liquid phases, hereby forming granules. Mutations within the TDP-43 proteins Glycine Rich Region (GRR) have recently been identified as associates that can contribute to various neurodegenerative diseases, with the most notable and common NDD being ALS, about 10% of the mutations causing familial ALS are accredited with the TDP-43 protein This CTD is often reported to play important role in pathogenic behavior of TDP-43: RNPs granules could have a role in stress response, and thus, aging, or persistence stress could lead the LLPs to turn into irreversible Liquid Solid Phase separation, pathological aggregates notably found in ALS neurons. The CTD's disorganized structure can turn into a full fledged amyloid-like beta-sheet rich structure, causing it to adopt prion-like properties. Moreover, CTFs are a common marker in diseased neurons and are argued to be of high toxicity. However, notice is to be taken that some points are not always consensual. Indeed, due to its hydrophobic structure, TDP-43 can be hard to analyze, and parts of it remain somewhat vague. Precise sites of phosphorylation, methylation, or even binding are still a bit elusive. == Function ==

TDP-43 is a transcriptional repressor that binds to chromosomally integrated TAR DNA and represses HIV-1 transcription. In addition, this protein regulates alternate splicing of the CFTR gene. In particular, TDP-43 is a splicing factor binding to the intron8/exon9 junction of the CFTR gene and to the intron2/exon3 region of the apoA-II gene. A similar pseudogene is present on chromosome 20. TDP-43 has been shown to bind both DNA and RNA and have multiple functions in transcriptional repression, pre-mRNA splicing and translational regulation. Recent work has characterized the transcriptome-wide binding sites revealing that thousands of RNAs are bound by TDP-43 in neurons. TDP-43 was originally identified as a transcriptional repressor that binds to chromosomally integrated trans-activation response element (TAR) DNA and represses HIV-1 transcription. In spinal motor neurons TDP-43 has also been shown in humans to be a low molecular weight neurofilament (hNFL) mRNA-binding protein. It has been demonstrated that zinc ions are able to induce aggregation of endogenous TDP-43 in cells. Moreover, zinc could bind to RNA binding domain of TDP-43 and induce the formation of amyloid-like aggregates in vitro. Consistently, Zn2+ binds a short peptide (residues 256–264) from the C-terminus of TDP-43's RRM2 domain with an association constant of ~1.6×105 M−1, supporting a specific Zn2+ site that may regulate nucleic-acid binding. DNA repair TDP-43 protein is a key element of the non-homologous end joining (NHEJ) enzymatic pathway that repairs DNA double-strand breaks (DSBs) in pluripotent stem cell-derived motor neurons. TDP-43 is rapidly recruited to DSBs where it acts as a scaffold for the further recruitment of the XRCC4-DNA ligase protein complex that then acts to seal the DNA breaks. In TDP-43 depleted human neural stem cell-derived motor neurons, as well as in sporadic ALS patients' spinal cord specimens there is significant DSB accumulation and reduced levels of NHEJ. == Clinical significance ==

A hyper-phosphorylated, ubiquitinated and cleaved form of TDP-43—known as pathologic TDP43—is the major disease protein in ubiquitin-positive, tau-, and alpha-synuclein-negative frontotemporal dementia (FTLD-TDP, previously referred to as FTLD-U) and in amyotrophic lateral sclerosis (ALS). Elevated levels of the TDP-43 protein have also been identified in individuals diagnosed with chronic traumatic encephalopathy, and has also been associated with ALS leading to the inference that athletes who have experienced multiple concussions and other types of head injury are at an increased risk for both encephalopathy and motor neuron disease (ALS). Abnormalities of TDP-43 also occur in an important subset of Alzheimer's disease patients, correlating with clinical and neuropathologic features indexes. Misfolded TDP-43 is found in the brains of older adults over age 85 with limbic-predominant age-related TDP-43 encephalopathy, (LATE), a form of dementia. New monoclonal antibodies, 2G11 and 2H1, have been developed to specify different TDP-43 inclusion types that occur across neurodegenerative diseases, without relying on hyper-phosphorylated epitopes. These antibodies were raised against an epitope within the RRM2 domain (amino acid residues 198–216). In particular, the TDP-43 mutants M337V and Q331K are being studied for their roles in ALS. While the aberrant mislocalization and cytoplasmic aggregation of TDP-43 characterizes FTLD with TDP-43 pathology (FTLD-TDP), recent work suggests the amyloid fibrils found in human FTLD-TDP brains are composed of transmembrane lysosomal protein TMEM106b rather than TDP-43. Cytoplasmic TDP-43 pathology is the dominant histopathological feature of multisystem proteinopathy. The N-terminal domain, which contributes importantly to the aggregation of the C-terminal region, has a novel structure with two negatively charged loops. A recent study has demonstrated that cellular stress can trigger the abnormal cytoplasmic mislocalisation of TDP-43 in spinal motor neurons in vivo, providing insight into how TDP-43 pathology may develop in sporadic ALS patients. ==Figures==