Structural determinants of the cellular localization and shuttling of TDP-43

Abstract

TDP-43 (also known as TARDBP) regulates different processes of gene expression, including transcription and splicing, through RNA and DNA binding. Moreover, recent reports have shown that the protein interacts with the 3′UTRs of specific mRNAs. The aberrant cellular distribution and aggregation of TDP-43 were recently reported in neurodegenerative diseases, namely frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). A detailed description of the determinants for cellular localization has yet to emerge, including information on how the known functions of TDP-43 and cellular targeting affect each other. We provide the first experimental evidence that TDP-43 continuously shuttles between nucleus and cytoplasm in a transcription-dependent manner. Furthermore, we investigate the role of the functional TDP-43 domains in determining cellular targeting through a combination of immunofluorescence and biochemical fractionation methods. Our analyses indicate that the C-terminus is essential for solubility and cellular localization, because its deletion results in the formation of large nuclear and cytoplasmic aggregates. Disruption of the RNA-recognition domain required for RNA and DNA binding, however, alters nuclear distribution by decreasing TDP-43 presence in the nucleoplasm. Our findings suggest that TDP-43 solubility and localization are particularly sensitive to disruptions that extend beyond the newly found nuclear localization signal and depend on a combination of factors that are closely connected to the functional properties of this protein.

TDP-43 regulatesβ-adducin(Add2) transcript stability

“…The siRNA sequences were as follows: TDP-43 siRNA 5 0 -GCAAAGCCAAGAUGAGCCU-3 0 , Luciferase siRNA 5 0 -UAAGGCUAUGAAGAGAUAC-3 0 . 43,44 RNA preparation, synthesis of cDNA, PCRs RNA preparation. Total cellular RNA was isolated with TRI REAGENT (Sigma) following manufacturer's instructions.…”

Mitochondrial Dysfunction and Decrease in Body Weight of a Transgenic Knock-in Mouse Model for TDP-43

“…TDP-43 is a widely expressed heterogeneous nuclear ribonucleoprotein with two RNA recognition motifs and high affinity to single-stranded nucleic acids (9 -14). Although predominantly located in the nucleus, TDP-43 shuttles between the nucleus and the cytoplasm under normal conditions (15). In the nucleus, TDP-43 is involved in microRNA biogenesis (16) and splicing regulation, where it causes exon inclusion or exclusion, depending on the number and localization of its UGenriched target sequences (17)(18)(19)(20)(21)(22).…”

A role for calpain-dependent cleavage of TDP-43 in amyotrophic lateral sclerosis pathology

“…Abnormal TDP-43 fragments are present in the central nervous system (CNS) of patients with ALS and frontotemporal lobar degeneration with TDP-43-positive inclusions (FTLD-TDP) 9,20 and it is believed that abnormally cleaved, aggregation-prone TDP-43 fragments serve as seeds for cytoplasmic inclusions and that normal, nuclear TDP-43 is sequestered by these inclusions when it shuttles between the nucleus and the cytoplasm 21 . However, the mechanism by which TDP-43 is cleaved into aggregation-prone fragments is unknown.…”