Inhibition of Aggregation of Mutant Huntingtin by Nucleic Acid Aptamers In Vitro and in a Yeast Model of Huntington's Disease

Chaudhary, Rajeev; Patel, Kinjal; Patel, M.; Joshi, Radha H; Roy, Ipsita

doi:10.1038/mt.2015.157

Cited by 39 publications

(69 citation statements)

References 53 publications

(79 reference statements)

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“…Also, no protein was retained on the cellulose acetate membrane in this case (Figure 2d), indicating that 20Q‐htt did not form aggregates. This confirmed the specificity of thioflavin T for fibrillar aggregates 26,28 and also showed that aggregation occurred due to the expanded polyQ stretch and not due to the presence of GST tag.…”

Section: Resultssupporting

confidence: 74%

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Stabilization of elongated polyglutamine tracts by a helical peptide derived from N‐terminal huntingtin

Sethi

Roy

2020

IUBMB Life

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In Huntington's disease, the length of the polyglutamine tract in the mutant protein correlates positively with the formation of aggregates and disease symptoms and severity of the disease. Some disease‐modifying factors exist. However, no organized study has been carried out to investigate the effect of polyglutamine length in the mutant protein on the efficacy of a therapeutic strategy. We had shown earlier that the helical peptide arising out of the N‐terminal stretch of normal huntingtin is able to inhibit aggregation of a number of proteins, including luciferase, α‐synuclein, p53, and Rnq1. In this work, we show that polyglutamine stretches of differing lengths, namely 51Q, 72Q, and 103Q, form a mixture of aggregates at different rates, with the rate increasing in a polyQ length‐dependent manner. The helical peptide is able to inhibit the rate of aggregation. The extent of inhibition was different when measuring either total aggregation or only fibrillar aggregates, suggesting that the helical peptide with benign polyQ stretch alters the aggregation landscape of different elongated polyQ lengths differently. Our results suggest that designing a therapeutic approach to inhibit protein aggregation must take note of polyQ length of the protein.

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Section: Resultssupporting

confidence: 74%

“…Expression and purification of GST‐20Q‐htt and GST‐51Q‐htt was carried out as described earlier 25,26 …”

Section: Methodsmentioning

confidence: 99%

Stabilization of elongated polyglutamine tracts by a helical peptide derived from N‐terminal huntingtin

Sethi

Roy

2020

IUBMB Life

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“…The use of nucleic acid aptamers is one such approach that has shown preliminary promise as a therapeutic strategy. 34 Aptamers are relatively short single-stranded oligonucleotides (DNA or RNA) that assume specific, stable conformations and bind tightly and specifically to protein targets by shape complementarity. 35 , 36 , 37 They can act as a stabilizer of the target protein in two ways: (1) by sterically hindering the unfolding of target protein monomer and (2) by inhibiting protein-protein interaction via electrostatic repulsion.…”

Section: Introductionmentioning

confidence: 99%

RNA Aptamers Rescue Mitochondrial Dysfunction in a Yeast Model of Huntington’s Disease

Patel

Chaudhary

Roy

2018

Molecular Therapy - Nucleic Acids

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Huntington’s disease (HD) is associated with the misfolding and aggregation of mutant huntingtin harboring an elongated polyglutamine stretch at its N terminus. A distinguishing pathological hallmark of HD is mitochondrial dysfunction. Any strategy that can restore the integrity of the mitochondrial environment should have beneficial consequences for the disease. Specific RNA aptamers were selected that were able to inhibit aggregation of elongated polyglutamine stretch containing mutant huntingtin fragment (103Q-htt). They were successful in reducing the calcium overload, which leads to mitochondrial membrane depolarization in case of HD. In one case, the level of Ca2+ was restored to the level of cells not expressing 103Q-htt, suggesting complete recovery. The presence of aptamers was able to increase mitochondrial mass in cells expressing 103Q-htt, along with rescuing loss of mitochondrial genome. The oxidative damage to the proteome was prevented, which led to increased viability of cells, as monitored by flow cytometry. Thus, the presence of aptamers was able to inhibit aggregation of mutant huntingtin fragment and restore mitochondrial dysfunction in the HD cell model, confirming the advantage of the strategy in a disease-relevant parameter.

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“…Interestingly, a peak at lower particle size (28.61 nm) was also observed, indicating the presence of oligomers. It has been reported that shorter polyQ stretches of benign length do not aggregate at all or do so at much longer time scales than the ones used here . Thus, it may be inferred that in the presence of FLAG‐N17‐25Q‐EGFP, luciferase forms low molecular weight oligomers and the aggregation of luciferase is suppressed.…”

Section: Resultsmentioning

confidence: 62%

Does N‐terminal huntingtin function as a ‘holdase’ for inhibiting cellular protein aggregation?

et al. 2018

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Proteolytic cleavage of huntingtin gives rise to N-terminal fragments. While the role of truncated mutant huntingtin is described in Huntington's disease (HD) pathogenesis, the function of N-terminal wild-type protein is less studied. The yeast model of HD is generated by the presence of FLAG tag and absence of polyproline tract as flanking sequences of the elongated polyglutamine stretch. We show that the same sequence derived from wild-type huntingtin exon1 is able to inhibit the aggregation of proteins in vitro and in yeast cells. It is able to stabilize client proteins as varied as luciferase, α-synuclein, and p53 in a soluble but non-native state. This is somewhat similar to the 'holdase' function of small heat shock proteins and 'nonchaperone proteins' which are able to stabilize partially unfolded client proteins in a nonspecific manner, slowing down their aggregation. Mutagenesis studies show this property to be localized at the N17 domain preceding the polyglutamine tract. Distortion of this ordered segment, either by deletion of this segment or mutation of a single residue (L4A), leads to decreased stability and increased aggregation of client proteins. It is interesting to note that the helical conformation of the N17 domain is also essential for aggregation of the N-terminal mutant protein. Our results provide evidence for a novel function for the amphipathic helix derived from exon1 of wild-type huntingtin.

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Inhibition of Aggregation of Mutant Huntingtin by Nucleic Acid Aptamers In Vitro and in a Yeast Model of Huntington's Disease

Cited by 39 publications

References 53 publications

Stabilization of elongated polyglutamine tracts by a helical peptide derived from N‐terminal huntingtin

Stabilization of elongated polyglutamine tracts by a helical peptide derived from N‐terminal huntingtin

RNA Aptamers Rescue Mitochondrial Dysfunction in a Yeast Model of Huntington’s Disease

Does N‐terminal huntingtin function as a ‘holdase’ for inhibiting cellular protein aggregation?

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