Involvement of CjMDR1, a plant multidrug-resistance-type ATP-binding cassette protein, in alkaloid transport in Coptis japonica

Abstract

Alkaloids comprise one of the largest groups of plant secondary metabolites. Berberine, a benzylisoquinoline alkaloid, is preferentially accumulated in the rhizome of Coptis japonica, a ranunculaceous plant, whereas gene expression for berberine biosynthetic enzymes has been observed specifically in root tissues, which suggests that berberine synthesized in the root is transported to the rhizome, where there is high accumulation. We recently isolated a cDNA encoding a multidrug-resistance protein (MDR)-type ATP-binding cassette (ABC) transporter (Cjmdr1) from berberineproducing cultured C. japonica cells, which is highly expressed in the rhizome. Functional analysis of Cjmdr1 by using a Xenopus oocyte expression system showed that CjMDR1 transported berberine in an inward direction, resulting in a higher accumulation of berberine in Cjmdr1-injected oocytes than in the control. Typical inhibitors of ABC proteins, such as vanadate, nifedipine, and glibenclamide, as well as ATP depletion, clearly inhibited this CjMDR1-dependent berberine uptake, suggesting that CjMDR1 functioned as an ABC transporter. Conventional membrane separation methods showed that CjMDR1 was localized in the plasma membrane of C. japonica cells. In situ hybridization indicated that Cjmdr1 mRNA was expressed preferentially in xylem tissues of the rhizome. These findings strongly suggest that CjMDR1 is involved in the translocation of berberine from the root to the rhizome.

Tobacco nicotine uptake permease (NUP1) affects alkaloid metabolism

“…In tobacco, the plasma membrane-localized NUP1 moved apoplastic nicotine into the cytoplasm. Similarly, the C. japonica CjMDR1 mobilizes the alkaloid berberine from the apoplast into the cytoplasm (28,41). However, NUP1 and CjMDR1 differ in how the alkaloid uptake is energized.…”

“…However, NUP1 and CjMDR1 differ in how the alkaloid uptake is energized. Based on similarity to AtPUP1 (29,31), NUP1 most likely uses proton symport for the uptake of nicotine, whereas CjMDR1 uses the hydrolysis of ATP to energize the transport of berberine into the cytoplasm (28). Both tobacco and C. japonica use the antiport of protons from the vacuole in conjunction with MATE-type membrane transporters to move cytoplasmic alkaloids into the vacuole (25,26,42).…”

“…One apoplastic alkaloid uptake transporter has been characterized. Coptis japonica multidrug resistance 1 (CjMDR1) is an ATPbinding cassette (ABC) plasma membrane protein responsible for the uptake of berberine into rhizome cells, where it accumulates to high levels (28). In this report, we describe a category of plasma membrane alkaloid transporter, called nicotine uptake permease 1 (NUP1), that was highly selective for nicotine uptake, and lines with reduced NUP1 expression levels resulted in altered nicotine metabolism, localization, and increased root growth.…”

Benzylisoquinoline alkaloid biosynthesis in opium poppy

“…Although the mechanism of translocation is unknown, the occurrence of plasmodesmata between opium poppy sieve elements and laticifers (Facchini and DeLuca 2008) suggests that both symplastic and apoplastic transport routes are possible. An ATP-binding cassette (ABC) transporter has been implicated in the uptake (Sakai et al 2002) and efflux (Terasaka et al 2003) of berberine in Coptis japonica, and the corresponding gene has been isolated (Shitan et al 2003(Shitan et al , 2012. However, it is not known whether an ABC transporter performs a similar function in opium poppy.…”

MDR‐like ABC transporter AtPGP4 is involved in auxin‐mediated lateral root and root hair development

“…In summary, both root transport and yeast complementation data point to an in vivo uptake of IAA over the plasma membrane. Uptake by an ABC MDR-like ABC transporter is an unusual event and has to our knowledge only been demonstrated once: CjMDR1 from Coptis japonica has been shown to catalyze the primary active uptake of the alkaloid berberine [27]. Interestingly, AtPGP4 shares (beside to its closest homologue AtPGP21) highest homology with CjMDR1 (65%) and in a phylogenetic tree AtPGP4/AtPGP21 and CjMDR1 cluster together on a subcluster that is clearly distinct from the one that is characterized by AtPGP1 and AtPGP19 (results not shown).…”