WILD-TYPE AND MUTANT HUNTINGTINS FUNCTION IN VESICLE TRAFFICKING IN THE SECRETORY AND ENDOCYTIC PATHWAYS

Huntingtin is a cytoplasmic protein that is found in neurons and somat ic cells. In patients with Huntington's disease (HD), the NH2-terminal region of huntingtin has an expanded polyglutamine tract. An abnormal protein interaction by mutant huntingtin has been proposed as a mecha nism for HD pathogenesis. Huntingtin associates with vesicle membranes and interacts with proteins involved in vesicle trafficking. It is un clear where along vesicle transport pathways wildtype and mutant hunti ngtins are found and whether polyglutamine expansion affects this loca lization. To distinguish wild-type and mutant huntingtin, fibroblasts from normals and HD patients with two mutant alleles (homozygotes) mer e examined. Immunofluorescence confocal microscopy showed that mutant huntingtin localized with clathrin in membranes of the trans Golgi net work and in clathrin;coated and noncoated endosomal vesicles in the cy toplasm and along plasma membranes. Separation of organelles in Nycode nz gradients showed that in normal and HD homozygote patient cells, hu ntingtin was present in membrane fractions enriched in clathrin. Simil ar results were obtained in fibroblasts from heterozyote juvenile HD p atients who had a highly expanded polyglutamine tract in the HD allele . Western blot analysis of membrane fractions from rat brain showed th at mild-type huntingtin was present in fractions that contained purifi ed clathrin-coated membranes or a mixture of clathrin-coated and nonco ated membranes. Electron microscopy of huntingtin immunoreactivity in rat brain revealed labeling along dendritic plasma membranes in associ ation with clathrin-coated pits and clusters of noncoated endosomal ve sicles 40-60 nm in diameter. These data suggest that wild-type and mut ant huntingtin can influence vesicle transport in the secretory and en docytic pathways through associations with clathrin-coated vesicles. ( C) 1998 Academic Press.