Two B-type ATP-binding cassette (ABC) transporters localize to the plasma membrane in Thalictrum minus

Alkaloids play important roles in plant defenses against herbivores and some alkaloids have medicinal uses. Medicinal alkaloids can be purified from plant tissues or produced axenically in cell culture systems. In culture, cells generally accumulate these toxic metabolites in the vacuole; however, treatment with benzyladenine (BA) induces cultured Thalictrum minus cells to produce the isoquinoline alkaloid berberine, which they release into the medium. A previous biochemical analysis suggested that B-type ATP-binding cassette (ABC) transporters participate in berberine efflux from cultured T. minus cells. In this study, we isolated full-length cDNAs of two novel B-type ABC transporter genes from T. minus, Tmabcb1 and Tmabcb2. The encoded transporters show significant amino acid sequence identity to the Coptis japonica berberine transporters CjABCB1 and CjABCB2. Real-time quantitative reverse transcription PCR analyses showed that BA induces an increase in Tmabcb1 and Tmabcb2 mRNA levels in cultured cells. Membrane separation and immunoblot analyses indicated that these proteins localize to the plasma membrane in T. minus cells. These data suggest that TmABCB1 and TmABCB2 participate in berberine transport in T. minus cells.