The polar transport of auxin is critical for development and growth of plants, and understanding how the directional movement of this hormone is controlled is a major challenge in plant biology. In this issue of New Phytologist, Rigas et al. (pp. 1130–1141) demonstrate that the polar localization of PIN-FORMED (PIN) proteins, which are responsible for the polar transport of auxin, requires the activity of TINY ROOT HAIR1 (TRH1), a putative plasma membrane potassium transporter.
‘The discovery that TRH1 is required for the localization of PIN1 is an important step forward in understanding how polar auxin transport is regulated.’
Auxin, synthesized in the shoot system, moves to the root tip through the stele and endodermis at the centre of the root. This ‘downward’ flow is facilitated by PIN proteins that transport auxin across the plasma membrane, and which are located on the face of the cell that is nearest to the root tip. PIN proteins accumulate in this plasma membrane domain in auxin-transporting cells and are excluded from other membrane domains on the sides or the domain facing the shoot.
The auxin transported across the plasma membrane may then be taken up by the neighbouring cell before being ‘effluxed’ across the membrane nearest the root tip again. Auxin is thus passed from cell to cell from the shoot to the root tip. Once at the root tip some auxin is channelled away from the tip in the outer layers of the root by a similar mechanism, except this time PIN proteins are located at the opposite end of the cell, nearest the shoot. This root-tip directed movement through the central regions of the root, and shootward movement through the peripheral tissues, has been likened to an upside-down fountain.
The accumulation of PIN proteins to specific membrane domains is thus central to polar auxin transport and identifying proteins that are required for the polar localization of PIN proteins in cells is an active field of enquiry. The paper by Rigas et al. shows that TRH1 activity is required for the polar localization of PIN1 proteins in roots.
This flux of auxin in and around the root tip coordinates development and growth responses including root-hair growth and gravitropism (the growth of roots relative to the gravity vector). Plants that lack TRH1 function have defects in both root-hair development and gravitropism, which can be suppressed by auxin treatment suggesting that TRH1 is active in a process involving auxin (Fig. 1a). Furthermore there is evidence that TRH1 plays a role in auxin transport itself. Nevertheless the precise mechanism of TRH1's involvement in auxin transport has remained elusive.
Rigas et al. set out to determine the way in which TRH1 facilitates auxin transport. They predicted that if TRH1 is required for the transport of auxin towards the root tip, then auxin would back up along this route in trh1 mutants. Such a blockage would be expected to result in higher levels of auxin behind the root tip. Indeed the levels of auxin in these regions were higher in trh1 mutants than in wild type. They then demonstrated that the levels of the auxin-responsive promoter DR5, were higher in this region of the root. Together these data are consistent with the hypothesis that TRH1 is required for the movement of auxin from shoot to root.
Given that the movement of auxin to the root tip is defective in trh1 mutants the authors hypothesized that TRH1 would either itself be asymmetrically localized in auxin-transporting cells or that it would control the polar localization of PIN1. They first demonstrated that TRH1:YFP is not uniformally distributed around the plasma membranes of cells; it is located on the plasma membrane domain nearest the root tip. This corresponds to the site of PIN1 protein accumulation, that is, TRH1 and PIN1 co-localize (Fig. 1b). To ascertain if TRH1 has a role in the regulation of PIN1 they determined the location of PIN1 in trh1 mutants. PIN1 is present in the membrane domain nearest the root tip in wild type but PIN1 is delocalized in the trh1 mutants – it accumulates in the plasma membrane nearest the root tip as in wild type but it also accumulates at the opposite end of the cell, closest to the shoot. TRH1 is therefore required for the localization of PIN1 protein to the plasma membrane domain nearest the root tip in stele cells.
Rigas et al. hypothesized that the delocalization of PIN1 proteins in trh1 mutants was sufficient to account for the defective root-hair and gravitropism phenotypes observed in trh1 mutants. To test the hypothesis, plants were generated in which PIN1 accumulated throughout the plasma membrane of stele cells (such delocalized PIN1 accumulation is found in plants that accumulate large amounts of PIN1 proteins). This can be done experimentally by expressing PIN1 at high levels using the cauliflower mosaic virus 35S (35S) promoter. Just as predicted, the roots of 35S:PIN1 roots were phenotypically similar to trh1 mutant roots – few root hairs developed and gravitropic responses were defective (Fig. 1). To independently verify this conclusion the authors treated roots with hypaphorine, a fungal drug that blocks the localization of PIN proteins to restricted domains in the plasma membrane. Hypaphorine treatment both reduced root-hair growth and caused gravitropic defects. Together these data support the hypothesis that delocalization of PIN1 proteins causes defects in root-hair growth and gravitropism, and is consistent with a model in which TRH1 regulates PIN1 localization during wild type development.
The paper by Rigas et al. demonstrates that TRH1 is a regulator of PIN1 localization and polar auxin transport. The discovery that TRH1 is required for the localization of PIN1 is an important step forward in understanding how polar auxin transport is regulated. Nevertheless, this discovery raises many new questions. Given that PIN1 and TRH1 co-localize, it is possible that these proteins physically interact in the target membrane. If they do, is this interaction required for localization of both? It is nevertheless formally possible that these proteins control auxin transport independently. TRH1 may itself modulate auxin transport, and this would in turn modulate PIN localization. Intriguingly TRH1 is likely to be a cation transporter. Is TRH1 cation transport activity required for its own localization and the localization of PIN1? Characterization of new trh1 alleles may provide us with the answers to these questions.