Inactivation of hnRNP K by expanded intronic AUUCU repeat induces apoptosis via translocation of PKCdelta to mitochondria in spinocerebellar ataxia 10.

Misti C. White, Rui Gao, Weidong Xu, Santi M. Mandal, Jung G. Lim, Tapas Hazra, Maki Wakamiya, Sharon F. Edwards, Salmo Raskin, Hélio A.G. Teive, Huda Y. Zoghbi, Partha Sarkar, Tetsuo Ashizawa

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Abstract

We have identified a large expansion of an ATTCT repeat within intron 9 of ATXN10 on chromosome 22q13.31 as the genetic mutation of spinocerebellar ataxia type 10 (SCA10). Our subsequent studies indicated that neither a gain nor a loss of function of ataxin 10 is likely the major pathogenic mechanism of SCA10. Here, using SCA10 cells, and transfected cells and transgenic mouse brain expressing expanded intronic AUUCU repeats as disease models, we show evidence for a key pathogenic molecular mechanism of SCA10. First, we studied the fate of the mutant repeat RNA by in situ hybridization. A Cy3-(AGAAU)(10) riboprobe detected expanded AUUCU repeats aggregated in foci in SCA10 cells. Pull-down and co-immunoprecipitation data suggested that expanded AUUCU repeats within the spliced intronic sequence strongly bind to hnRNP K. Co-localization of hnRNP K and the AUUCU repeat aggregates in the transgenic mouse brain and transfected cells confirmed this interaction. To examine the impact of this interaction on hnRNP K function, we performed RT-PCR analysis of a splicing-regulatory target of hnRNP K, and found diminished hnRNP K activity in SCA10 cells. Cells expressing expanded AUUCU repeats underwent apoptosis, which accompanied massive translocation of PKCdelta to mitochondria and activation of caspase 3. Importantly, siRNA-mediated hnRNP K deficiency also caused the same apoptotic event in otherwise normal cells, and over-expression of hnRNP K rescued cells expressing expanded AUUCU repeats from apoptosis, suggesting that the loss of function of hnRNP K plays a key role in cell death of SCA10. These results suggest that the expanded AUUCU-repeat in the intronic RNA undergoes normal transcription and splicing, but causes apoptosis via an activation cascade involving a loss of hnRNP K activities, massive translocation of PKCdelta to mitochondria, and caspase 3 activation.

Original languageEnglish (US)
JournalPLoS Genetics
Volume6
Issue number6
DOIs
StatePublished - 2010

Fingerprint

Heterogeneous-Nuclear Ribonucleoprotein K
apoptosis
mitochondrion
translocation
inactivation
Mitochondria
mitochondria
Apoptosis
RNA
brain
cells
chromosome
mutation
caspase-3
Caspase 3
Transgenic Mice
genetically modified organisms
loss
Spinocerebellar Ataxia 10
disease models

ASJC Scopus subject areas

  • Genetics
  • Molecular Biology
  • Ecology, Evolution, Behavior and Systematics
  • Cancer Research
  • Genetics(clinical)

Cite this

Inactivation of hnRNP K by expanded intronic AUUCU repeat induces apoptosis via translocation of PKCdelta to mitochondria in spinocerebellar ataxia 10. / White, Misti C.; Gao, Rui; Xu, Weidong; Mandal, Santi M.; Lim, Jung G.; Hazra, Tapas; Wakamiya, Maki; Edwards, Sharon F.; Raskin, Salmo; Teive, Hélio A.G.; Zoghbi, Huda Y.; Sarkar, Partha; Ashizawa, Tetsuo.

In: PLoS Genetics, Vol. 6, No. 6, 2010.

Research output: Contribution to journalArticle

White, Misti C. ; Gao, Rui ; Xu, Weidong ; Mandal, Santi M. ; Lim, Jung G. ; Hazra, Tapas ; Wakamiya, Maki ; Edwards, Sharon F. ; Raskin, Salmo ; Teive, Hélio A.G. ; Zoghbi, Huda Y. ; Sarkar, Partha ; Ashizawa, Tetsuo. / Inactivation of hnRNP K by expanded intronic AUUCU repeat induces apoptosis via translocation of PKCdelta to mitochondria in spinocerebellar ataxia 10. In: PLoS Genetics. 2010 ; Vol. 6, No. 6.
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abstract = "We have identified a large expansion of an ATTCT repeat within intron 9 of ATXN10 on chromosome 22q13.31 as the genetic mutation of spinocerebellar ataxia type 10 (SCA10). Our subsequent studies indicated that neither a gain nor a loss of function of ataxin 10 is likely the major pathogenic mechanism of SCA10. Here, using SCA10 cells, and transfected cells and transgenic mouse brain expressing expanded intronic AUUCU repeats as disease models, we show evidence for a key pathogenic molecular mechanism of SCA10. First, we studied the fate of the mutant repeat RNA by in situ hybridization. A Cy3-(AGAAU)(10) riboprobe detected expanded AUUCU repeats aggregated in foci in SCA10 cells. Pull-down and co-immunoprecipitation data suggested that expanded AUUCU repeats within the spliced intronic sequence strongly bind to hnRNP K. Co-localization of hnRNP K and the AUUCU repeat aggregates in the transgenic mouse brain and transfected cells confirmed this interaction. To examine the impact of this interaction on hnRNP K function, we performed RT-PCR analysis of a splicing-regulatory target of hnRNP K, and found diminished hnRNP K activity in SCA10 cells. Cells expressing expanded AUUCU repeats underwent apoptosis, which accompanied massive translocation of PKCdelta to mitochondria and activation of caspase 3. Importantly, siRNA-mediated hnRNP K deficiency also caused the same apoptotic event in otherwise normal cells, and over-expression of hnRNP K rescued cells expressing expanded AUUCU repeats from apoptosis, suggesting that the loss of function of hnRNP K plays a key role in cell death of SCA10. These results suggest that the expanded AUUCU-repeat in the intronic RNA undergoes normal transcription and splicing, but causes apoptosis via an activation cascade involving a loss of hnRNP K activities, massive translocation of PKCdelta to mitochondria, and caspase 3 activation.",
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AU - White, Misti C.

AU - Gao, Rui

AU - Xu, Weidong

AU - Mandal, Santi M.

AU - Lim, Jung G.

AU - Hazra, Tapas

AU - Wakamiya, Maki

AU - Edwards, Sharon F.

AU - Raskin, Salmo

AU - Teive, Hélio A.G.

AU - Zoghbi, Huda Y.

AU - Sarkar, Partha

AU - Ashizawa, Tetsuo

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