Alkoxy-auxins Are Selective Inhibitors of Auxin Transport Mediated by PIN, ABCB, and AUX1 Transporters

Abstract

Polar auxin movement is a primary regulator of programmed and plastic plant development. Auxin transport is highly regulated at the cellular level and is mediated by coordinated transport activity of plasma membrane-localized PIN, ABCB, and AUX1/LAX transporters. The activity of these transporters has been extensively analyzed using a combination of pharmacological inhibitors, synthetic auxins, and knock-out mutants in Arabidopsis. However, efforts to analyze auxin-dependent growth in other species that are less tractable to genetic manipulation require more selective inhibitors than are currently available. In this report, we characterize the inhibitory activity of 5-alkoxy derivatives of indole 3-acetic acid and 7-alkoxy derivatives of naphthalene 1-acetic acid, finding that the hexyloxy and benzyloxy derivatives act as potent inhibitors of auxin action in plants. These alkoxy-auxin analogs inhibit polar auxin transport and tropic responses associated with asymmetric auxin distribution in Arabidopsis and maize. The alkoxy-auxin analogs inhibit auxin transport mediated by AUX1, PIN, and ABCB proteins expressed in yeast. However, these analogs did not inhibit or activate SCF TIR1 auxin signaling and had no effect on the subcellular trafficking of PIN proteins. Together these results indicate that alkoxy-auxins are inactive auxin analogs for auxin signaling, but are recognized by PIN, ABCB, and AUX1 auxin transport proteins. Alkoxyauxins are powerful new tools for analyses of auxin-dependent development.

Auxin transport sites are visualized in planta using fluorescent auxin analogs

“…1A) (18). These alkoxy-auxin analogs are inactive at auxin receptors but are recognized as auxin by auxin transporters.…”

“…These alkoxy-auxin analogs are inactive at auxin receptors but are recognized as auxin by auxin transporters. A structureactivity analysis revealed that auxin transporters ignore the alkoxychain substructure of auxin analogs (18). Based on our previous findings, we synthesized 5-fluorescently labeled IAAs and 7-fluorescently labeled NAAs as analogs of IAA and NAA, respectively (Fig.…”

“…S1). The artificial auxins NAA and Bz-NAA were efficiently exported by efflux transporters but were not imported by the AUX1 influx symporter, suggesting that fluorescent NAA analogs can be used to visualize efflux transport via PINs and ABCBs (18). The BODIPY and NBD fluorescent dyes were introduced into 5-hydroxy-IAA and 7-hydroxy-NAA using various alkyl linkers (SI Appendix, Fig.…”

Block of ATP-Binding Cassette B19 Ion Channel Activity by 5-Nitro-2-(3-Phenylpropylamino)-Benzoic Acid Impairs Polar Auxin Transport and Root Gravitropism

“…It blocks polar auxin transport, binds to at least ABCB1, and interferes with ABCB1-TWD1 interactions (Kim et al, 2010). The structures of 2-[4-(diethylamino)-2-hydroxybenzoyl]benzoic acid and IAA derivatives that also block auxin transport at least in part by interfering with ABCB transport function (Tsuda et al, 2011) are shown in Figure 7.…”

“…Mutations can create complicating pleiotropic effects by inhibiting the process throughout development, while inhibitors can be used to impose an effect at a specific time. 1-Naphthylphthalamic acid (NPA) is the most commonly used inhibitor of polar auxin transport (Katekar and Geissler, 1980), but others are being discovered (Rojas-Pierce et al, 2007;Kim et al, 2010;Tsuda et al, 2011). Inhibitors are especially useful when their targets are well defined, which would be the case if NPPB blocked ABCB19 and induced its expression as hypothesized.…”