title: Phosphorylation regulates proteasomal-mediated degradation and solubility of TAR DNA binding protein-43 C-terminal fragments.
abstract: BACKGROUND
  Inclusions of TAR DNA binding protein-43 (TDP-43) are the defining histopathological feature of several neurodegenerative diseases collectively referred to as TDP-43 proteinopathies. These diseases are characterized by the presence of cellular aggregates composed of abnormally phosphorylated, N-terminally truncated and ubiquitinated TDP-43 in the spinal cord and/or brain. Recent studies indicate that C-terminal fragments of TDP-43 are aggregation-prone and induce cytotoxicity. However, little is known regarding the pathways responsible for the degradation of these fragments and how their phosphorylation contributes to the pathogenesis of disease.
  
  
  RESULTS
  Herein, we established a human neuroblastoma cell line (M17D3) that conditionally expresses an enhanced green fluorescent protein (GFP)-tagged caspase-cleaved C-terminal TDP-43 fragment (GFP-TDP220-414). We report that expression of this fragment within cells leads to a time-dependent formation of inclusions that are immunoreactive for both ubiquitin and phosphorylated TDP-43, thus recapitulating pathological hallmarks of TDP-43 proteinopathies. Phosphorylation of GFP-TDP220-414 renders it resistant to degradation and enhances its accumulation into insoluble aggregates. Nonetheless, GFP-TDP220-414 inclusions are reversible and can be cleared through the ubiquitin proteasome system. Moreover, both Hsp70 and Hsp90 bind to GFP-TDP220-414 and regulate its degradation.
  
  
  CONCLUSIONS
  Our data indicates that inclusions formed from TDP-43 C-terminal fragments are reversible. Given that TDP-43 inclusions have been shown to confer toxicity, our findings have important therapeutic implications and suggest that modulating the phosphorylation state of TDP-43 C-terminal fragments may be a promising therapeutic strategy to clear TDP-43 inclusions.
authors.affiliation: Department of Neuroscience, Mayo Clinic, 4500 San Pablo Road, Jacksonville, Florida 32224, USA. petrucelli.leonard@mayo.edu.
authors.forename: Yong-Jie
authors.initials: YJ
authors.lastname: Zhang
authors.forename: Tania F
authors.initials: TF
authors.lastname: Gendron
authors.forename: Ya-Fei
authors.initials: YF
authors.lastname: Xu
authors.forename: Li-Wen
authors.initials: LW
authors.lastname: Ko
authors.forename: Shu-Hui
authors.initials: SH
authors.lastname: Yen
authors.forename: Leonard
authors.initials: L
authors.lastname: Petrucelli
references.citation: FASEB J. 2006 Apr;20(6):753-5
references.pmid: 16464956
references.citation: J Neurosci Res. 2010 Mar;88(4):784-97
references.pmid: 19798749
references.citation: J Neurochem. 2004 Feb;88(3):545-53
references.pmid: 14720204
references.citation: Neurosci Lett. 2010 Jan 18;469(1):112-6
references.pmid: 19944744
references.citation: J Alzheimers Dis. 2004 Dec;6(6):605-22; discussion 673-81
references.pmid: 15665401
references.citation: Neuron. 2003 May 8;38(3):375-87
references.pmid: 12741986
references.citation: EMBO J. 2000 Nov 1;19(21):5720-8
references.pmid: 11060023
references.citation: Science. 2006 Oct 6;314(5796):130-3
references.pmid: 17023659
references.citation: Proc Natl Acad Sci U S A. 2009 May 5;106(18):7607-12
references.pmid: 19383787
references.citation: Ann Neurol. 2008 Jul;64(1):60-70
references.pmid: 18546284
references.citation: Proc Natl Acad Sci U S A. 2008 Apr 29;105(17):6439-44
references.pmid: 18434538
references.citation: J Biol Chem. 2009 Mar 27;284(13):8516-24
references.pmid: 19164285
references.citation: Mol Neurodegener. 2009 Mar 11;4:13
references.pmid: 19284597
references.citation: Biochem Biophys Res Commun. 2006 Dec 22;351(3):602-11
references.pmid: 17084815
references.citation: Cell. 2008 Aug 22;134(4):679-91
references.pmid: 18724940
references.citation: J Clin Invest. 2007 Mar;117(3):648-58
references.pmid: 17304350
references.citation: J Neurosci. 2007 Sep 26;27(39):10530-4
references.pmid: 17898224
references.citation: J Biol Chem. 2009 Oct 2;284(40):27416-24
references.pmid: 19651785
references.citation: Acta Neuropathol. 2006 Sep;112(3):237-51
references.pmid: 16845533
references.citation: J Neurochem. 2009 Aug;110(3):1082-94
references.pmid: 19522733
references.citation: J Biol Chem. 2005 Mar 25;280(12):11635-40
references.pmid: 15664989
references.citation: J Biol Chem. 2009 Jul 24;284(30):20329-39
references.pmid: 19465477
references.citation: Hum Mol Genet. 2009 Sep 15;18(18):3353-64
references.pmid: 19515851
references.citation: J Biol Chem. 2009 Oct 9;284(41):28253-28262
references.pmid: 19679666
references.citation: J Biomed Biotechnol. 2006;2006(3):74539
references.pmid: 17047313