Grapevine MATE-Type Proteins Act as Vacuolar H+-Dependent Acylated Anthocyanin Transporters    

Abstract

In grapevine (Vitis vinifera), anthocyanins are responsible for most of the red, blue, and purple pigmentation found in the skin of berries. In cells, anthocyanins are synthesized in the cytoplasm and accumulated into the vacuole. However, little is known about the transport of these compounds through the tonoplast. Recently, the sequencing of the grapevine genome allowed us to identify genes encoding proteins with high sequence similarity to the Multidrug And Toxic Extrusion (MATE) family. Among them, we selected two genes as anthocyanin transporter candidates and named them anthoMATE1 (AM1) and AM3. The expression of both genes was mainly fruit specific and concomitant with the accumulation of anthocyanin pigment. Subcellular localization assays in grapevine hairy roots stably transformed with AM1∷ or AM3∷green fluorescent protein fusion protein revealed that AM1 and AM3 are primarily localized to the tonoplast. Yeast vesicles expressing anthoMATEs transported acylated anthocyanins in the presence of MgATP. Inhibitor studies demonstrated that AM1 and AM3 proteins act in vitro as vacuolar H+-dependent acylated anthocyanin transporters. By contrast, under our experimental conditions, anthoMATEs could not transport malvidin 3-O-glucoside or cyanidin 3-O-glucoside, suggesting that the acyl conjugation was essential for the uptake. Taken together, these results provide evidence that in vitro the two grapevine AM1 and AM3 proteins mediate specifically acylated anthocyanin transport.

ABCC1, an ATP Binding Cassette Protein from Grape Berry, Transports Anthocyanidin 3-O-Glucosides

“…Collected berries were immediately frozen in liquid nitrogen and stored at 280°C before use. Plant material (roots, shoots, leaves, and berries) were also collected from grapevine plants grown in Sup Agro-Institut National de la Recherche Agronomique as previously described by Gomez et al (2009).…”

“…Concerning the transporter-mediated model, biochemical, molecular, and genetic evidence support the involvement of both multidrug and toxic extrusion (MATE) and the ATP binding cassette (ABC) proteins in the transport of anthocyanins (Goodman et al, 2004;Marinova et al, 2007b;Gomez et al, 2009;Zhao and Dixon, 2009;Zhao et al, 2011). MATE transporters comprise a large family of transporters, widely distributed in all living organisms.…”

Vacuolar Transport of Abscisic Acid Glucosyl Ester Is Mediated by ATP-Binding Cassette and Proton-Antiport Mechanisms in Arabidopsis

“…The total protein concentration in microsomal extract was quantified using the Bradford assay (Bio-Rad; with BSA as a standard). The intactness of the microsomal preparations was assessed using the 9-amino-6-chloro-2-methoxyacridine dye fluorescence quenching method described by Gomez et al (2009).…”

Grape Berry Vacuole: A Complex and Heterogeneous Membrane System Specialized in the Accumulation of Solutes

“…Recently, and due to the sequencing of the grapevine genome (Jaillon et al 2007), two grapevine proteins belonging to the multidrug and toxic extrusion (MATE) family, anthoMATE1 (AM1) and anthoMATE3 (AM3), have been implicated in the mediated transport of specifically acylated anthocyanin (Gomez et al 2009). Vacuolar sequestration of anthocyanins is an important process for cell survival because anthocyanins are believed to be toxic (reviewed by Boss and Davies 2009).…”

“…A number of studies involving the identification of several enzymes, transcriptional regulators of proanthocyanidin biosynthesis, and transporters have illustrated the movement of proanthocyanidin precursors into the vacuole (Terrier et al 2009). Tannins are accumulated in specific vacuoles (tannin vacuoles) and act as deterrents to herbivores (Boss and Davies 2009, Jackson 2008, Conn et al 2003, Gomez et al 2009 Nonflavonoid � ++ +++ Plant defense Includes caftaric and coumaric acids, benzoic and cinnamic acids; nonflavonoid compounds and tannins are the most accumulated compounds in grape seeds; synthesis maintained throughout ripening (Kramling and Singleton 1969, De 2000, Montealegre et al 2006 a Relative abundance of the compound in each tissue ranges from very abundant (+++) to absent (�).…”