Analysis of the subcellular localization of huntingtin with a set of rabbit polyclonal antibodies in cultured mammalian cells of neuronal origin: comparison with the distribution of huntingtin in Huntington's disease autopsybrain

Huntington's disease (HD) is a neurodegenerative disorder with a midlife on set. The disease is caused by expansion of a CAG (glutamine) repeat within the coding region of the HD gene. The molecular mechanism by which the muta ted protein causes this disease is still unclear. To study the protein we h ave generated a set of rabbit polyclonal antibodies raised against differen t segments of the N-terminal, central and C-terminal parts of the protein. The polyclonal antibodies were affinity purified and characterized in ELISA and Western blotting experiments. All antibodies can react with mouse and human proteins. The specificity of these antibodies is underscored by their recognition of huntingtin with different repeat sizes in extracts prepared from patient-derived lymphoblasts. The antibodies were used in immunofluor escence experiments to study the subcellular localization of huntingtin in mouse neuroblastoma N1E-115 cells. The results indicate that most huntingti n is present in the cytoplasm, whereas a minor fraction is present in the n ucleus. On differentiation of the N1E-115 cells in vitro, the subcellular d istribution of huntingtin does not change significantly. These results sugg est that full-length huntingtin with a normal repeat length can be detected in the nucleus of cycling and non-cycling cultured mammalian cells of neur onal origin. However, in HD autopsy brain the huntingtin-containing neurona l intranuclear inclusions can be detected only with antibodies raised again st the N-terminus of huntingtin. Thus several forms of huntingtin display t he propensity for nuclear localization, possibly with different functional consequences.