Spliceosome integrity is defective in the motor neuron diseases ALS and SMA.
Bottom Line: We also show that in ALS, Gems are lost, U snRNA levels are up-regulated and spliceosomal U snRNPs abnormally and extensively accumulate in motor neuron nuclei, but not in the temporal lobe of FTLD with TDP-43 pathology.This aberrant accumulation of U snRNAs in ALS motor neurons is in direct contrast to SMA motor neurons, which show reduced amounts of U snRNAs, while both have defects in the spliceosome.These findings indicate that a profound loss of spliceosome integrity is a critical mechanism common to neurodegeneration in ALS and SMA, and may explain cell-type specific vulnerability of motor neurons.
Affiliation: Laboratory for Motor Neuron Disease, RIKEN Brain Science Institute, Wako, Saitama, Japan. firstname.lastname@example.orgShow MeSH
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Mentions: The association and localization of TDP-43, FUS/TLS and SMN in Gems imply a functional convergence of three proteins. SMN is well known to assist in assembly of U snRNPs, which is central to splicing, in the cytoplasm and to recruit U snRNPs into the nucleus (Pellizzoni et al, 2002). In SMA mice and SMN-depleted cells, the levels of U snRNAs and components of U snRNPs are unbalanced, resulting in aberrant splicing (Gabanella et al, 2007; Zhang et al, 2008). Therefore, we hypothesized that TDP-43 might have a function in U snRNP biogenesis and alterations in U snRNPs may be also responsible for ALS. To test this hypothesis, we first analysed if TDP-43 associated with U snRNPs. TDP-43 distribution was similar to U snRNPs, which were marked by the anti-dimethylated Sm proteins antibody (Y12), and both TDP-43 and U snRNPs were concentrated to same nuclear bodies in Hela cells and primary cultured mouse hippocampal neurons (Fig 4A, arrows), suggesting the interaction of TDP-43 and snRNPs. Since C-terminus of TDP-43 is required for TDP-43-containing foci in nuclei (Fig 2B, Supporting Information Fig S3C), we identified proteins interacting with C-terminus of TDP-43. Comparison of proteins immunoprecipitated with wild type FLAG-tagged TDP-43 or deletion mutant of C-terminal domain, followed by LC–MS/MS, revealed many proteins associated with a TDP-43 C-terminus including U snRNP components such as PRPF3 (Supporting Information Fig S4A and B). The association of TDP-43 with U snRNP components PRPF3 and U1-70K was confirmed by IP-Western blotting (Supporting Information Fig S4C). To investigate the association between U snRNPs and TDP-43, U snRNPs were immunoprecipitated with the anti-Sm proteins antibody (Y12) (Supporting Information Fig S4D). Subsequent immunoblotting confirmed that TDP-43 was co-immunoprecipitated with U snRNPs although at a relatively low level. These results suggest a possible involvement of TDP-43 in maintaining the integrity of U snRNPs.
Affiliation: Laboratory for Motor Neuron Disease, RIKEN Brain Science Institute, Wako, Saitama, Japan. email@example.com