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

Citation
J. Ko et al., New anti-huntingtin monoclonal antibodies: Implications for huntingtin conformation and its binding proteins, BRAIN RES B, 56(3-4), 2001, pp. 319-329
Citations number
31
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Neurosciences & Behavoir
Journal title
BRAIN RESEARCH BULLETIN
ISSN journal
0361-9230 → ACNP
Volume
56
Issue
3-4
Year of publication
2001
Pages
319 - 329
Database
ISI
SICI code
0361-9230(200110/11)56:3-4<319:NAMAIF>2.0.ZU;2-S
Abstract
We produced eight anti-huntingtin (Htt) monoclonal antibodies (mAbs), sever al of which have novel binding patterns. Peptide array epitope mapping show s that mAbs MW1-6 specifically bind the polyQ domain of Htt exon 1. On West ern blots of extracts from mutant Htt knock-in mouse brain and Huntington's disease lymphoblastoma cell lines, MW1-5 all strongly prefer to bind to th e expanded polyQ repeat form of Htt, displaying no detectable binding to no rmal Htt. These results suggest that the polyQ domain can assume different conformations that are distinguishable by mAbs. This idea is supported by i mmunohistochemistry with wild type (WT) and mutant Htt transgenic mouse (R6 ) brains. Despite sharing the same epitope and binding preferences on Weste rn blots, MW1-5 display distinct staining patterns. MW1 shows punctate cyto plasmic and neuropil staining, while MW2-5 strongly stain the neuronal Golg i complex. MW6, in contrast, stains neuronal somas and neuropil. In additio n, despite their preference for mutant Htt on blots, none of these mAbs sho w enhanced staining of R6 brains over WT, and show no binding of the Htt-co ntaining nuclear inclusions in R6 brains. This suggests that in its various subcellular locations, the polyQ domain of Htt either takes on different c onformations and/or is differentially occluded by Htt binding proteins. In contrast to MW1-6, MW7, and 8 can differentiate transgenic from WT mice by staining nuclear inclusions in R6/2 brain; MW8 displays no detectable stain ing in WT brain and stains only inclusions in R6/2 brain. Epitope mapping r eveals that MW7 and 8 specifically bind the polyp domains and amino acids 8 3-90, respectively. As with MW1-6, the epitopes for MW7 and 8 are different ially available in the various subcellular compartments where Htt is found. (C) 2001 Elsevier Science Inc.