Identification of substrates of two plant pleiotropic drug resistance transporters, NtPDR1 from Nicotiana tabacum and MpPDR1 from Mentha x piperita

Leaf trichomes are particularly rich in the production of secondary metabolites that are involved in the plant protection against pathogens. Some of these metabolites are also toxic for the cells that produce them and have therefore to be actively transported out of the cell. Very little is known concerning their transport although in some cases huge concentration differences are observed between cellular compartments.\nTwo Pleiotropic Drug Resistance (PDR)-type ABC transporters expressed in trichomes were characterized. We hypothesized that they extrude metabolites involved in the plant response to (a)biotic stress. NtPDR1, expressed in Nicotiana tabacum glandular trichomes, is probably involved in the plant defense by transporting antimicrobial diterpenes. However, direct transport has not been demonstrated yet. The second transporter, MpPDR1, expressed in Mentha x piperita glandular trichomes, is presumably involved in monoterpene transport. The aim of the project was to identify the substrates of these PDRs.\nMpPDR1 was first expressed in the yeast, Saccharomyces cerevisiae. However, this system was not appropriate since MpPDR1 was mislocalized. Therefore, both transporters were expressed in N. tabacum BY2 suspension cells. In situ immunolocalization showed plasma membrane localization of both PDRs as in the plant. Toxicity assays with putative substrates showed that NtPDR1-expressing lines were more resistant to the diterpenes sclareol, manool, abietic acid and dehydroabietic acid, suggesting their export out of the cell. Transport assays confirmed some of them and also identified the diterpene cembrene as a substrate. However, other molecules such as mono- and sesquiterpenes were not transported. On the other hand, M. x piperita abundant monoterpenes were not identified as MpPDR1 substrates. Nevertheless, cembrene was exported by MpPDR1, indicating that this transporter is also involved in diterpene transport.