Structural Biochemistry/TPD-43

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TPD-43[edit]

The protein 43 kDa TAR DNA binding protein (TDP-43) was first discovered to be involved in neurodegeneration. TPD-43 was discovered to have involvement in amyotrophic lateral sclerosis (ALS) and frontotemporal labor degeneration (FTLD) in 2006, which lead an increase laboratories that studied this protein. This is because unusual TPD-43 aggregation is the main focus of many neuronal diseases that are known as TPD-43 proteinopathies. A new and promising research area had been opened up after TPD-43 was discovered to be involved in neurodegeneration, and findings of other proteins FUS/TLS and C9orf72. These discoveries also opened up the field of RNA binding proteins in neuroscience. But there it is difficult to study TPD-43, because of large amount of process that it is involved in. TPD-43 is a protein that is essential to a cell's life cycle.

TPD-43 RNA Binding[edit]

TPD-43 is in a family of nuclear factors named hnRNPs. The ability of TPD-43 to bind RNA on specific sequences in a single-stranded behavior. This ability comes from the sixty amino acid residue motifs that are folded in a conserved 3D structure, known as RNA recognition motifs (RRMs). This ability of TPD-43 to bind to RNA is important for RNA processes, especially in alternative splicing. TPD-43 binding to repeats of UG RNA sequences can silence splicing. This can been confirmed in vivo in some labs using in vivo crosslinking and immunoprecipitation-sequencing. Some of these studies have shown that TPD-43 can bind to other conserved sequence motifs, but the function of these binding sites are not well known. UG RNA sequence repeats are common in the human genome and are found mostly in introns and 3' regions that are not translated. TPD-43 also controls neurofilament mRNA stability with the help of other proteins in the ALS pathology.

An extended binding region, TDPBR, of TPD-43 was found in the TPD-43 mRNA 3' UTR. The TDPBR has a few non-UG sequences that are important for autoregulation of TPD-43 mRNA levels. Low levels of nuclear TPD-43 causes the uses of the most efficient polyA1 instead of other choices like polyA2 to polyA4. On the other hand, high levels of nuclear TPD-43 causes less optimal splice sites to be used instead, which leads to rapid degradation of the mRNA. This makes a feedback loop that the cell uses to keep the concentration of TPD-43 leveled. TPD-43 aggregation can occur if the levels are not kept constant. If this happens, TPD-43 aggregates in the cell nucleus and cytoplasm will reduce the amount of free nuclear TPD-43. Then the 3' UTR TDPBR sensor will read protein level drops and increase TPD-43 production. This is bad, because this cycle would lead to cell stress and eventually, death. While high levels of TPD-43 can be bad, low levels and can also have serious effects on expression levels and RNA transcripts.

TPD-43 Protein Interactions[edit]

Another important aspect of TPD-43 is with its ability to bind proteins that help with its RNA processing abilities and aggregation properties. Proteins like hnRNP A1 and A2 are necessary for inhibiting splicing. Another important protein is FUS/TLS, which regulates the expression levels of histone deacetylase 6. All these interactions show that TPD-43 is a very flexible protein in its interactions proteins and RNA. This is important when searching for therapeutic uses of TPD-43 in splicing targets.

TDP-43 Properties of Aggregation[edit]

It is accepted that the C-terminal is the biggest reason that TPD-43 aggregates. Modifications in TPD-43 such as C-terminal fragments are found in neurons of patients with ALS and FTLD. De novo nuclear cleavage of TPD-43 makes C-terminal fragments, which has been shown to lead to aggregation of TPD-43. Hyperphosphorylation of TPD-43 could be protective to neurons, in contrast to aggregation promotion effects. However, further studies are needed before this can be concluded. TPD-43 aggregation has also been observed when nuclear transport proteins, like karyopherin beta or cellular apoptosis susceptibility proteins, are knocked down. An external factor that could increase TPD-43 aggregation is their interaction with aggregates of polygluatmine like Ataxin-2, or extracellular kinase (ERK1/2) inhibtion. Overexpression of p62 protein and USP14 inhibition go against TPD-43 aggregation. This indicates a possible relationship with autophagosome system in aggregate resolution. Bad autophagosome functioning could contribute to promoting disease growth.

TPD-43 is brought to stress granules after cell stress. TPD-43 controls levels of stress granule factors for formation and support. Some of these factors are GTPase activating protein and TIA-1 binding protein. Long period of cell stress can lead to stress granule aggregates.

TPD-43 in FTLD and ALS[edit]

Research has shown that TPD-43 can act as biochemical marker in FTLD and ALS. It is important to indentify these markers before there is too much neuronal damage from FTLD and ALS. Biomarkers are important when judging the effects of therapeutic techniques. Research has been focused on finding pathological modifications of proteins (including: amyloid beta, tau protein, prion protein, alpha-synuclein, and TPD-43 ) that are important for neurodegeneration. The tissue samples that have been studied for abnormal TPD-43 levels are cerebrospinal fluid, blood plasma, circulating lymphonocytes, and skeletal muscle in people with ALS. These studies can be difficult, especially in cerebrospinal fluid and blood plasma. One method that could be easier is to use TPD-43 solubility tests for proteins from peripheral blood mononuclear cells. Using these tissues makes it easier to compare mRNA expression and polyadenylation levels with normal cells.