The assembly of alpha-HL was shown to rapidly progress upon its interaction with Caveolin-1. Treatment of A431 cells with alpha-HL has resulted in clustering of Caveolin-1 at cell-cell contacts. Consistent with this observation, alpha-HL mutants devoid of assembly property have not induced the clustering of Caveolin-1. While cholesterol depletion of A431 cells completely arrests the assembly of alpha-HL, chelation of membrane cholesterol results in its retarded assembly. Interestingly, HT29 cells, with low Caveolin-1 levels, are resistant to a-HL attack. Clustering of Caveolin-1, as seen in case of A431 cells, was readily observed in case of HT29 cells transfected with Caveolin-1 construct, thus overexpressing the full length Caveolin-1, upon alpha-HL treatment. A model was constructed to visualize the interactions between a-HL and Caveolin-1 which suggests that facile penetration of alpha-HL's P-barrel might occur through protein-protein interactions with the surrounding 7 alpha-helices of Caveolin-1. (C) 2004 Elsevier Inc. All rights reserved.