Flavonoids Redirect PIN-mediated Polar Auxin Fluxes during Root Gravitropic Responses

Abstract

The rate, polarity, and symmetry of the flow of the plant hormone auxin are determined by the polar cellular localization of PIN-FORMED (PIN) auxin efflux carriers. Flavonoids, a class of secondary plant metabolites, have been suspected to modulate auxin transport and tropic responses. Nevertheless, the identity of specific flavonoid compounds involved and their molecular function and targets in vivo are essentially unknown. Here we show that the root elongation zone of agravitropic pin2/eir1/ wav6/agr1 has an altered pattern and amount of flavonol glycosides. Application of nanomolar concentrations of flavonols to pin2 roots is sufficient to partially restore root gravitropism. By employing a quantitative cell biological approach, we demonstrate that flavonoids partially restore the formation of lateral auxin gradients in the absence of PIN2. Chemical complementation by flavonoids correlates with an asymmetric distribution of the PIN1 protein. pin2 complementation probably does not result from inhibition of auxin efflux, as supply of the auxin transport inhibitor N-1-naphthylphthalamic acid failed to restore pin2 gravitropism. We propose that flavonoids promote asymmetric PIN shifts during gravity stimulation, thus redirecting basipetal auxin streams necessary for root bending.The plant hormone auxin (3-indolyl acetic acid, IAA) 5 controls virtually all plant developmental and physiological processes. In roots, the differential growth response associated with gravity stimulation (gravitropism) occurs in the elongation zone (1, 2) and is a result of the asymmetric distribution of auxin to the lower side of epidermal cells (3). In these tissues accumulating auxin, cell elongation is inhibited and the root tip bends downwards. This cell-to-cell or polar auxin transport (PAT) is determined by the asymmetric cellular localization of auxin inand efflux components of the ABCB/PGP/MDR, AUX1/LAX, and PIN-FORMED (PIN) family (4 -7). Although ABCBs are apparently involved in long-range auxin transport and movements of auxin out of apical regions (8 -10), AUX1 and PIN2/ EIR1/WAV6AGR1 have been demonstrated to channel auxin from the lateral root cap basipetally to the expanding epidermal cells (11-13).The regulation of auxin transport during root gravitropic responses is still largely unclear. Among various possible mechanisms, the localized synthesis and directed transport of flavonoids, plant-specific phenylpropanoid compounds, have been shown to modulate the rate of the gravity response (14,15). A number of lines of experimentation have suggested that flavonoids may act as non-essential auxin transport inhibitors (16 -20). This is mainly based on the finding that flavonoids displace binding of synthetic auxin transport inhibitors, like N-1-naphthylphthalamic acid (NPA), a herbicide (Naptalam), in vitro (31, 50 -52). Moreover, roots of transparent testa (tt) Arabidopsis mutant with manipulated flavonoid levels exhibit altered gravitropic curvature and auxin transport, which are restored to the wild-type level by exogenous...

Identification of an ABCB/P-glycoprotein-specific Inhibitor of Auxin Transport by Chemical Genomics

“…A similar effect was found also for hypocotyl elongation of light-grown seedlings. NPA shows, in agreement with previous reports on roots (38), a stimulating and inhibitory result upon hypocotyl elongation under light at nanomolar and micromolar concentrations, respectively. Such a biphasic behavior was not found for BUM in the concentration range used (0 -20 M), suggesting, despite widely overlapping effects, a different mode of action or targets (Fig.…”

“…However, based on gravitropism assays (supplemental Fig. S2) and in contrast to what was recently found for NPA (11,38), abcb1 roots were significantly less affected by BUM: 35.3% (percentage occurrence of 60, 90, and 120°bending between inhibitor and solvent control (see "Experimental Procedures")) of abcb1 roots bent efficiently on BUM but only 26.8% of the corresponding wild type. Even higher resistance was found for pin2 root gravitropism (supplemental Fig.…”

“…S2). Interestingly, the ecotype Col revealed higher sensitivities (77% inhibition) compared with the ecotype Ws (58% inhibition) not found for NPA (11,38). Similarly to lateral root formation, 0.1 M BUM (supplemental Fig.…”

“…Similarly to lateral root formation, 0.1 M BUM (supplemental Fig. S2) was more efficient than 5 M NPA assayed in both ecotypes in parallel (not shown) (11,38). Recently, single loss-of-function roots, pin2, abcb1, or abcb19, were shown to be NPA-sensitive using gravitropism assays (11,38).…”

cis-Cinnamic Acid Is a Novel, Natural Auxin Efflux Inhibitor That Promotes Lateral Root Formation

“…Intriguingly, endogenous auxin transport inhibitors are scarce. Flavonols and flavonoids such as quercetin were considered to inhibit auxin transporters (Brown et al, 2001), although later work suggested that flavonoids also act by redirecting PIN efflux protein localization (Santelia et al, 2008). Certain flavonoid mutants display auxin-related defects (Buer et al, 2013) and an auxin-transport inhibiting activity was recently assigned to the flavonol glycoside kaempferol 3-O-rhamnoside-7-O-rhamnoside (Yin et al, 2014).…”