title: The regulation of N-terminal Huntingtin (Htt552) accumulation by Beclin1.
abstract: OBJECTIVE
  Huntingtin protein (Htt) was a neuropathological hallmark in human Huntington's Disease. The study aimed to investigate whether the macroautophagy regulator, Beclin1, was involved in the degradation of Htt.
  
  
  METHODS
  PC12 cells and primary cultured brain neurons of rats were examined. pDC316 adenovirus shuttle plasmid was used to mediate the expression of wild-type Htt-18Q-552 or mutant Htt-100Q-552 in PC12 cells. The expression of the autophagy-related proteins LC3 II and Beclin1, as well as the lysosome-associated enzymes Cathepsin B and L was evaluated using Western blotting. The locations of Beclin1 and Htt were observed with immunofluorescence and confocal microscope.
  
  
  RESULTS
  Htt552 expression increased the expression of LC3 II, Beclin1, cathepsin B and L in autophagy/lysosomal degradation pathway. Treatment with the autophagy inhibitor 3-MA or the proteasome inhibitors lactacystin and MG-132 increased Htt552 levels in PC12 cells infected with Ad-Htt-18Q-552 or Ad-Htt-100Q-552. The proteasome inhibitor caused a higher accumulation of Htt552-18Q than Htt552-100Q, and the autophagy inhibitor resulted in a higher accumulation of Htt552-100Q than Htt552-18Q. Similar results were observed in primary cultured neurons infected with adenovirus. In Htt552-expressing cells, Beclin1 was redistributed from the nucleus to the cytoplasm. Htt siRNA prevented Beclin1 redistribution in starvation conditions. Blockade of Beclin1 nuclear export by leptomycin B or Beclin1 deficiency caused by RNA interference induced the formation of mHtt552 aggregates.
  
  
  CONCLUSIONS
  Beclin1 regulates the accumulation of Htt via macroautophagy.
authors.affiliation: Department of Pharmacology, Laboratory of Aging and Nervous Diseases, Soochow University School of Pharmaceutical Science, Suzhou, China.
authors.forename: Jun-chao
authors.initials: JC
authors.lastname: Wu
authors.forename: Lin
authors.initials: L
authors.lastname: Qi
authors.forename: Yan
authors.initials: Y
authors.lastname: Wang
authors.forename: Kimberly B
authors.initials: KB
authors.lastname: Kegel
authors.forename: Jennifer
authors.initials: J
authors.lastname: Yoder
authors.forename: Marian
authors.initials: M
authors.lastname: Difiglia
authors.forename: Zheng-hong
authors.initials: ZH
authors.lastname: Qin
authors.forename: Fang
authors.initials: F
authors.lastname: Lin
mesh_terms: D021581:Active Transport, Cell Nucleus; D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D001343:Autophagy; D000071186:Beclin-1; D001921:Brain / Q000166:cytology / Q000378:metabolism / Q000473:pathology; D002478:Cells, Cultured; D006801:Humans; D000071058:Huntingtin Protein; D006816:Huntington Disease / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism* / Q000473:pathology; D009687:Nuclear Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D016716:PC12 Cells; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D015854:Up-Regulation
chemical_list: D051017:Apoptosis Regulatory Proteins; D000071186:Beclin-1; C512829:Becn1 protein, rat; C499524:Htt protein, rat; D000071058:Huntingtin Protein; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D034741:RNA, Small Interfering