Effects of Intracellular Expression of Anti-Huntingtin Antibodies of Various Specificities on Mutant Huntingtin Aggregation and Toxicity

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 99; no. 2; pp. 1002 - 1007
• Main Authors: Khoshnan, Ali, Ko, Jan, Patterson, Paul H.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 22.01.2002; National Acad Sciences; The National Academy of Sciences
• Subjects: Aggregation; Antibodies; Antibodies, Monoclonal - genetics; Antibodies, Monoclonal - immunology; Antibody Specificity; Apoptosis; Binding Sites, Antibody - genetics; Biological Sciences; Cell aggregates; Cell death; Cell Line; Cloning, Molecular; Exons; Humans; huntingtin; Huntingtin Protein; Huntington disease; Huntington Disease - etiology; Immunoglobulin Variable Region - genetics; Macromolecular Substances; Mutation; Nerve Tissue Proteins - chemistry; Nerve Tissue Proteins - genetics; Nerve Tissue Proteins - immunology; Nerve Tissue Proteins - toxicity; Neurological disorders; Neurons; Nuclear Proteins - chemistry; Nuclear Proteins - genetics; Nuclear Proteins - immunology; Nuclear Proteins - toxicity; PC12 cells; Peptides - chemistry; Proteins; Single-Chain Antibodies; Transfection
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.022631799

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.