MicroRNA pathways Modulate Polyglutamine-induced Neurodegeneration

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MicroRNA pathways Modulate Polyglutamine-induced
Neurodegeneration

• Julide Bilen, Nan Liu, Barrington G. Burnett,
  Randall N. Pitman, and Nancy M. Borris
• Molecular Cell, October 6, 2006
• Christine Marciniak
• February 21, 2008

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Background Human polyQ diseases MicroRNAs

• Expansion in CAG repeats
• Dominant toxicity ? Neuronal dysfunction loss
• Pathogenic protein accumulates in nuclear
  inclusions
• Some hallmarks of programmed cell-death seen
• (Condensation of nucleaus cytoplasm)
• Spinocerebeller ataxia type 3 (SCA3)
• miRNAs shown to regulate programmed cell-death
  genes in Drosophilia
• Do miRNAs play a role in neurodegeneration?

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Ataxin-3 Degeneration in the fly eye

• Weak (Q61) or partial (Q78 or Q84) Poly-Q
  induced degeneration
• Loss of Dicer-1 (or R3D1) leads to enhancement
  of Poly-Q induced degeneration
• (miRNA pathway affected)
• Loss of Dicer-1 (or R3D1) ALONE does not lead to
  degeneration
• Loss of Dicer-2 does not affect Poly-Q induced
  degeneration
• (siRNA pathway not affected)

External Eyes
Hoechst Staining Retinal Thickness
External Eyes Retinal Thickness
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Ataxin-3 Toxicity in Human Cells
Control siRNA or siRNA to dicer Short poly-Q
repeat (Q29)
Significant increase in cell death in
dicer-knockdown treated cells with
expansion Significant mitigation of cell death
in similar cells also treated with purified small
RNA fraction
Expanded poly-Q repeat (Q72)
No toxicity after 24hrs
Enhanced cell death with no Dicer
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miRNA bantam modulates Ataxin-3 toxicity
Genomic region of ban
Psuedopupil Assay Neurol Loss enhanced with loss
of ban
Suppression of Poly-Q toxicity by overexpression
of ban
External fly eye and Internal retinal sections
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ban miRNA suppresses degeneration downstream of
polyQ toxicity
Immunostaining in 7-day flies for stress-induced
Hsp-70 Endogenous stress response not altered by
ban
4-hour vs. 7-day flies Diffuse
Accumulated in nuclear inclusions polyQ protein
Western immunoblot for stress-induced
Hsp-70 Endogenous stress response not altered by
ban
7-day flies Coexpression ban
has no effect human Hsp-70 reduces accumulation
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Main conclusions

• If miRNAs are gone, more degeneration is observed
• Knockdown of dicer observed in polyQ disease
  SCA3 degenerated fly eye and cultured human cells
• Specific miRNAs are involved in the regulated
  pathway to modulate polyQ toxicity
• bantam miRNA
• NOT siRNAs! (Dicer-2 not affected in fly eye)