Dataset Information

Cellular localization and allele-selective inhibition of mutant huntingtin protein by peptide nucleic acid oligomers containing the fluorescent nucleobase [bis-o-(aminoethoxy)phenyl]pyrrolocytosine.

ABSTRACT: Peptide nucleic acid (PNA) is a successful DNA/RNA mimic. A major challenge for research is to invent chemically modified PNAs that retain the favorable properties of the parent compound while improving biological recognition. Here, we test modified PNAs containing [bis-o-(aminoethoxy)phenyl]pyrrolocytosine bases designed to engage guanine with an additional hydrogen bond. We observe elevated melting temperatures, localization to cellular compartments, and allele-selective inhibition of mutant huntingtin protein expression.

SUBMITTER: Hu J

PROVIDER: S-EPMC2770837 | biostudies-literature | 2009 Nov

REPOSITORIES: biostudies-literature

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Publications

Cellular localization and allele-selective inhibition of mutant huntingtin protein by peptide nucleic acid oligomers containing the fluorescent nucleobase [bis-o-(aminoethoxy)phenyl]pyrrolocytosine.

Hu Jiaxin J Dodd David W DW Hudson Robert H E RH Corey David R DR

Bioorganic & medicinal chemistry letters 20090906 21

Peptide nucleic acid (PNA) is a successful DNA/RNA mimic. A major challenge for research is to invent chemically modified PNAs that retain the favorable properties of the parent compound while improving biological recognition. Here, we test modified PNAs containing [bis-o-(aminoethoxy)phenyl]pyrrolocytosine bases designed to engage guanine with an additional hydrogen bond. We observe elevated melting temperatures, localization to cellular compartments, and allele-selective inhibition of mutant h ...[more]

PMID: 19783436

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Project description:Expansion of the polyglutamine (polyQ) track of the Huntingtin (HTT) protein above 36 is associated with a sharply enhanced risk of Huntington's disease (HD). Although there is general agreement that HTT toxicity resides primarily in N-terminal fragments such as the HTT exon1 protein, there is no consensus on the nature of the physical states of HTT exon1 that are induced by polyQ expansion, nor on which of these states might be responsible for toxicity. One hypothesis is that polyQ expansion induces an alternative, toxic conformation in the HTT exon1 monomer. Alternative hypotheses posit that the toxic species is one of several possible aggregated states. Defining the nature of the toxic species is particularly challenging because of facile interconversion between physical states as well as challenges to identifying these states, especially in vivo. Here we describe the use of fluorescence correlation spectroscopy (FCS) to characterize the detailed time and repeat length dependent self-association of HTT exon1-like fragments both with chemically synthesized peptides in vitro and with cell-produced proteins in extracts and in living cells. We find that, in vitro, mutant HTT exon1 peptides engage in polyQ repeat length dependent dimer and tetramer formation, followed by time dependent formation of diffusible spherical and fibrillar oligomers and finally by larger, sedimentable amyloid fibrils. For expanded polyQ HTT exon1 expressed in PC12 cells, monomers are absent, with tetramers being the smallest molecular form detected, followed in the incubation time course by small, diffusible aggregates at 6-9 hours and larger, sedimentable aggregates that begin to build up at 12 hrs. In these cell cultures, significant nuclear DNA damage appears by 6 hours, followed at later times by caspase 3 induction, mitochondrial dysfunction, and cell death. Our data thus defines limits on the sizes and concentrations of different physical states of HTT exon1 along the reaction profile in the context of emerging cellular distress. The data provide some new candidates for the toxic species and some new reservations about more well-established candidates. Compared to other known markers of HTT toxicity, nuclear DNA damage appears to be a relatively early pathological event.

Dataset Information

Cellular localization and allele-selective inhibition of mutant huntingtin protein by peptide nucleic acid oligomers containing the fluorescent nucleobase [bis-o-(aminoethoxy)phenyl]pyrrolocytosine.

Publications

Cellular localization and allele-selective inhibition of mutant huntingtin protein by peptide nucleic acid oligomers containing the fluorescent nucleobase [bis-o-(aminoethoxy)phenyl]pyrrolocytosine.

Similar Datasets

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