New anti-huntingtin monoclonal antibodies: implications for huntingtin conformation and its binding proteins

Ko, Jan; Ou, Susan; Patterson, Paul H.

doi:10.1016/s0361-9230(01)00599-8

Cited by 170 publications

(192 citation statements)

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“…We performed additional imaging studies utilizing the monoclonal antibody MW8 to detect YFP-HTTex1Q79 inclusion bodies. This antibody preferentially recognizes insoluble HTT protein aggregates in cells but not monomers or small oligomers (Ko et al 2001). The quantification of YFP fluorescence and MW8 immunoreactivity yielded very similar results (Fig.…”

Section: Crmp1 Expression Levels Are Altered In Brains Of Hd Transgensupporting

confidence: 54%

Systematic interaction network filtering identifies CRMP1 as a novel suppressor of huntingtin misfolding and neurotoxicity

et al. 2015

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Assemblies of huntingtin (HTT) fragments with expanded polyglutamine (polyQ) tracts are a pathological hallmark of Huntington's disease (HD). The molecular mechanisms by which these structures are formed and cause neuronal dysfunction and toxicity are poorly understood. Here, we utilized available gene expression data sets of selected brain regions of HD patients and controls for systematic interaction network filtering in order to predict disease-relevant, brain region-specific HTT interaction partners. Starting from a large protein-protein interaction (PPI) data set, a step-by-step computational filtering strategy facilitated the generation of a focused PPI network that directly or indirectly connects 13 proteins potentially dysregulated in HD with the disease protein HTT. This network enabled the discovery of the neuron-specific protein CRMP1 that targets aggregation-prone, N-terminal HTT fragments and suppresses their spontaneous self-assembly into proteotoxic structures in various models of HD. Experimental validation indicates that our network filtering procedure provides a simple but powerful strategy to identify disease-relevant proteins that influence misfolding and aggregation of polyQ disease proteins.

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Section: Crmp1 Expression Levels Are Altered In Brains Of Hd Transgensupporting

confidence: 54%

Systematic interaction network filtering identifies CRMP1 as a novel suppressor of huntingtin misfolding and neurotoxicity

et al. 2015

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“…Moreover, interaction of some of the WW proteins with htt exon 1 is enhanced in htt with an expanded polyQ stretch (20,21). Interestingly, the MW7 mAb selectively stains intranuclear aggregates formed in the transgenic mouse model (27). This finding implies that expanded polyQ makes the proline-rich motifs more available to binding to WW-containing proteins or to MW7.…”

Section: Discussionmentioning

confidence: 94%

“…We previously generated eight mAbs that bind the htt epitopes polyQ, polyP, or the C terminus of exon 1 (27). Although MW1 stains htt in the cytoplasm, MW2 displays vesicular staining of htt.…”

Section: Resultsmentioning

confidence: 99%

“…Although MW1 stains htt in the cytoplasm, MW2 displays vesicular staining of htt. Interestingly, MW7 stains htt in the perinuclear region as well in the nuclear inclusions formed by mutant htt (27). To examine the effects of these mAbs on the biological activities of htt exon 1, we generated scFvs for MW1 and MW2, which recognize the polyQ domain, and MW7, which recognizes the polyP domains of htt exon 1.…”

Section: Resultsmentioning

confidence: 99%

“…In fact, intracellular expression of an Ab against the N terminus of htt was shown to inhibit aggregate formation induced in cultured cells by mutant htt, although quantitative results on inhibiting htt toxicity were not reported (26). We have generated eight mAbs (MW1-8) that recognize polyQ, polyproline (polyP), or a unique epitope near the C terminus of htt exon 1 (27). Here we report that intracellular expression of some of these mAbs as recombinant, singlechain variable region fragments (scFvs) targeted to different regions of htt exon 1 can either block or enhance aggregation as well as the cell death induced by a mutant htt.…”

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confidence: 99%

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Effects of intracellular expression of anti-huntingtin antibodies of various specificities on mutant huntingtin aggregation and toxicity

Khoshnan

Ko²,

Patterson³

2002

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

140

114

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We have generated eight mAbs (MW1-8) that bind the epitopes polyglutamine (polyQ), polyproline (polyP), or the C terminus of exon 1 in huntingtin (htt) protein. In the brains of Huntington's disease (HD) mouse models, the anti-polyQ mAbs bind to various cytoplasmic compartments, whereas the anti-polyP and anti-C terminus mAbs bind nuclear inclusions containing htt. To use these mAbs as intracellular perturbation agents, we have cloned and expressed the antigen-binding domains of three of the mAbs as single-chain variable region fragment Abs (scFvs). In 293 cells cotransfected with htt exon 1 containing an expanded polyQ domain, MW1, MW2, and MW7 scFvs colocalize with htt exon 1. Moreover, these scFvs coimmunoprecipitate with htt exon 1 in cell extracts. In perturbation experiments, MW7 scFv, recognizing the polyP domains of htt, significantly inhibits aggregation as well as the cell death induced by mutant htt protein. In contrast, MW1 and MW2 scFvs, recognizing the polyQ stretch, stimulate htt aggregation and apoptosis. Therefore, these anti-htt scFvs can be used to investigate the role of the polyP and polyQ domains in HD pathogenesis, and antibody binding to the polyP domain has potential therapeutic value in HD. H untington's disease (HD), a fatal neurodegenerative disorder, is caused by abnormal expansion of CAG repeats that translate into an extended polyglutamine (polyQ) stretch in exon 1 of the protein huntingtin (htt) (1). Mutant htt with Ͼ40 CAG repeats gains a toxic function and induces death in subpopulations of neurons in the striatum and cortex (2-4). A hallmark of HD and other polyQ diseases is the formation of insoluble protein aggregates in affected neurons (5, 6). A major component of the aggregates in HD is the N terminus exon 1 of mutant htt (2,(5)(6)(7)(8). Abnormal behavior and aggregate formation are also seen in transgenic mice expressing htt exon 1 with an expanded polyQ stretch (9-11).Neuronal death in HD has been variously attributed to polyQ toxicity, activation of caspases, interference with transcriptional machinery, and sequestration͞inactivation of wild-type htt and other important cellular factors (12-17). Several proteins that interact with exon 1 of htt have been identified (14,(18)(19)(20)(21)(22)(23), and although the function of these proteins in the etiology of HD is unclear, the transcriptional coactivator CREB-binding protein (CBP) as well as proteins with WW domains have been implicated in the HD pathology (18-21). Binding of htt to CBP has been shown to repress CBP-mediated gene expression (18,19). Moreover, ectopic expression of CBP appears to block httmediated toxicity, indicating that transcriptional dysregulation may contribute to HD pathogenesis (19,24). Several different WW-containing proteins have been shown to interact with proline-rich domains in the C terminus of htt exon 1 (20, 21). These interactors include spliceosomes (HYPA and HYPC) and transcription factors (HYPB), which appear to have a higher affinity for expanded polyQ htt (20). By using specific anti...

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Protein misfolding inside cells: The case of huntingtin and Huntington's disease

Hatters

2008

IUBMB Life

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SummaryHuntington's disease is one of the several neurodegenerative diseases caused by dominant mutations that expand the number of glutamine codons within an existing poly-glutamine (polyQ) repeat sequence of a gene. An expanded polyQ sequence in the huntingtin gene is known to cause the huntingtin protein to aggregate and form intracellular inclusions as disease progresses. However, the role that polyQ-induced aggregation plays in disease is yet to be fully determined. This review focuses on key questions remaining for how the expanded polyQ sequences affect the aggregation properties of the huntingtin protein and the corresponding effects on cellular machinery. The scope includes the technical challenges that remain for rigorously assessing the effects of aggregation on the cellular machinery.2008 IUBMB IUBMB Life, 60(11): 724-728, 2008

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New anti-huntingtin monoclonal antibodies: implications for huntingtin conformation and its binding proteins

Cited by 170 publications

References 28 publications

Systematic interaction network filtering identifies CRMP1 as a novel suppressor of huntingtin misfolding and neurotoxicity

Systematic interaction network filtering identifies CRMP1 as a novel suppressor of huntingtin misfolding and neurotoxicity

Effects of intracellular expression of anti-huntingtin antibodies of various specificities on mutant huntingtin aggregation and toxicity

Protein misfolding inside cells: The case of huntingtin and Huntington's disease

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