Transient efflux inhibition improves plant regeneration by natural auxins.

Plant genome editing and propagation are important tools in crop breeding and production. Both rely heavily on the development of efficient in vitro plant regeneration systems. Two prominent regeneration systems that are widely employed in crop production are somatic embryogenesis (SE) and de novo shoot regeneration. In many of the protocols for SE or shoot regeneration, explants are treated with the synthetic auxin analog 2,4-dichlorophenoxyacetic acid (2,4-D), since natural auxins, such as indole-3-acetic acid (IAA) or 4-chloroindole-3-acetic acid (4-Cl-IAA), are less effective or even fail to induce regeneration. Based on previous reports that 2,4-D, compared to endogenous auxins, is not effectively exported from plant cells, we investigated whether efflux inhibition of endogenous auxins could convert these auxins into efficient inducers of SE in Arabidopsis immature zygotic embryos (IZEs). We show that natural auxins and synthetic analogs thereof become efficient inducers of SE when their efflux is transiently inhibited by co-application of the auxin transport inhibitor naphthylphthalamic acid (NPA). Moreover, IZEs of auxin efflux mutants pin2 or abcb1 abcb19 show enhanced SE efficiency when treated with IAA or efflux-inhibited IAA, confirming that auxin efflux reduces the efficiency of Arabidopsis SE. Importantly, in contrast to the 2,4-D system, where only 50-60% of the embryos converted to seedlings, all SEs induced by transport-inhibited natural auxins converted to seedlings. Efflux-inhibited IAA, like 2,4-D, also efficiently induced SE from carrot suspension cells, whereas IAA alone could not, and efflux-inhibited 4-Cl-IAA significantly improved de novo shoot regeneration in Brassica napus. Our data provides new insights into the action of 2,4-D as an efficient inducer of plant regeneration but also shows that replacing this synthetic auxin for efflux-inhibited natural auxin significantly improves different types of plant regeneration, leading to a more synchronized and homogenous development of the regenerated plants.

Far-red light-enhanced apical dominance stimulates flower and fruit abortion in sweet pepper.

Far-red radiation affects many plant processes, including reproductive organ abortion. Our research aimed to determine the role of apical dominance in far-red light-induced flower and fruit abortion in sweet pepper (Capsicum annuum L.). We conducted several climate room experiments where plants were grown under white- or red-rich LED light, with or without additional far-red light. Additional far-red light enhanced apical dominance: it increased auxin levels in the apices of dominant shoots, and caused a greater difference in internode length and apical auxin levels between dominant and subordinate shoots. Additional far-red light stimulated fruit abortion in intact plants but not in decapitated plants, suggesting a crucial role of shoot apices in this effect. However, reducing basipetal auxin transport in the stems with N-1-naphthylphthalamic acid did not influence far-red light-stimulated fruit abortion, although auxin levels in the stem were largely reduced. Applying the synthetic auxin 1-naphthaleneacetic acid on decapitated apices did not influence fruit abortion. However, applying the auxin biosynthesis inhibitor yucasin to shoot apices reduced fruit abortion regardless of the light conditions, accompanied by slight shoot growth retardation. These findings suggest that the basipetal auxin stream does not mediate far-red light-stimulated fruit abortion. Far-red light-stimulated fruit abortion was associated with reduced sucrose accumulation and lower invertase activities in flowers. We suggest that under additional far-red light conditions, increased auxin levels in shoot apices promote fruit abortion probably through enhanced competition for assimilates between apices and flowers, which limits assimilate import into flowers.

Endogenous auxin maintains embryonic cell identity and promotes somatic embryo development in Arabidopsis.

Somatic embryogenesis (SE), or embryo development from in vitro cultured vegetative explants, can be induced in Arabidopsis by the synthetic auxin 2,4-dichlorophenoxyacetic acid (2,4-D) or by overexpression of specific transcription factors, such as AT-HOOK MOTIF NUCLEAR LOCALIZED 15 (AHL15). Here, we explored the role of endogenous auxin [indole-3-acetic acid (IAA)] during 2,4-D and AHL15-induced SE. Using the pWOX2:NLS-YFP reporter, we identified three distinct developmental stages for 2,4-D and AHL15-induced SE in Arabidopsis, with these being (i) acquisition of embryo identity; (ii) formation of pro-embryos; and (iii) somatic embryo patterning and development. The acquisition of embryo identity coincided with enhanced expression of the indole-3-pyruvic acid auxin biosynthesis YUCCA genes, resulting in an enhanced pDR5:GFP-reported auxin response in the embryo-forming tissues. Chemical inhibition of the indole-3-pyruvic acid pathway did not affect the acquisition of embryo identity, but significantly reduced or completely inhibited the formation of pro-embryos. Co-application of IAA with auxin biosynthesis inhibitors in the AHL15-induced SE system rescued differentiated somatic embryo formation, confirming that increased IAA levels are important during the last two stages of SE. Our analyses also showed that polar auxin transport, with AUXIN/LIKE-AUX influx and PIN-FORMED1 efflux carriers as important drivers, is required for the transition of embryonic cells to proembryos and, later, for correct cell fate specification and differentiation. Taken together, our results indicate that endogenous IAA biosynthesis and its polar transport are not required for the acquisition of embryo identity, but rather to maintain embryonic cell identity and for the formation of multicellular proembryos and their development into histodifferentiated embryos.

Structure-activity relationship of 2,4-D correlates auxinic activity with the induction of somatic embryogenesis in Arabidopsis thaliana.

2,4-dichlorophenoxyacetic acid (2,4-D) is a synthetic analogue of the plant hormone auxin that is commonly used in many in vitro plant regeneration systems, such as somatic embryogenesis (SE). Its effectiveness in inducing SE, compared to the natural auxin indole-3-acetic acid (IAA), has been attributed to the stress triggered by this compound rather than its auxinic activity. However, this hypothesis has never been thoroughly tested. Here we used a library of forty 2,4-D analogues to test the structure-activity relationship with respect to the capacity to induce SE and auxinic activity in Arabidopsis thaliana. Four analogues induced SE as effectively as 2,4-D and 13 analogues induced SE but were less effective. Based on root growth inhibition and auxin response reporter expression, the 2,4-D analogues were classified into different groups, ranging from very active to not active auxin analogues. A halogen at the 4-position of the aromatic ring was important for auxinic activity, whereas a halogen at the 3-position resulted in reduced activity. Moreover, a small substitution at the carboxylate chain was tolerated, as was extending the carboxylate chain with an even number of carbons. The auxinic activity of most 2,4-D analogues was consistent with their simulated TIR1-Aux/IAA coreceptor binding characteristics. A strong correlation was observed between SE induction efficiency and auxinic activity, which is in line with our observation that 2,4-D-induced SE and stress both require TIR1/AFB auxin co-receptor function. Our data indicate that the stress-related effects triggered by 2,4-D and considered important for SE induction are downstream of auxin signalling.

Local phytochrome signalling limits root growth in light by repressing auxin biosynthesis.

In nature, plant shoots are exposed to light whereas the roots grow in relative darkness. Surprisingly, many root studies rely on in vitro systems that leave the roots exposed to light whilst ignoring the possible effects of this light on root development. Here, we investigated how direct root illumination affects root growth and development in Arabidopsis and tomato. Our results show that in light-grown Arabidopsis roots, activation of local phytochrome A and B by far-red or red light inhibits respectively PHYTOCHROME INTERACTING FACTORS 1 or 4, resulting in decreased YUCCA4 and YUCCA6 expression. As a result, auxin levels in the root apex become suboptimal, ultimately resulting in reduced growth of light-grown roots. These findings highlight once more the importance of using in vitro systems where roots are grown in darkness for studies that focus on root system architecture. Moreover, we show that the response and components of this mechanism are conserved in tomato roots, thus indicating its importance for horticulture as well. Our findings open up new research possibilities to investigate the importance of light-induced root growth inhibition for plant development, possibly by exploring putative correlations with responses to other abiotic signals, such as temperature, gravity, touch, or salt stress.

Drought response in Arabidopsis displays synergistic coordination between stems and leaves.

The synergy between drought-responsive traits across different organs is crucial in the whole-plant mechanism influencing drought resilience. These organ interactions, however, are poorly understood, limiting our understanding of drought response strategies at the whole-plant level. Therefore, we need more integrative studies, especially on herbaceous species that represent many important food crops but remain underexplored in their drought response. We investigated inflorescence stems and rosette leaves of six Arabidopsis thaliana genotypes with contrasting drought tolerance, and combined anatomical observations with hydraulic measurements and gene expression studies to assess differences in drought response. The soc1ful double mutant was the most drought-tolerant genotype based on its synergistic combination of low stomatal conductance, largest stomatal safety margin, more stable leaf water potential during non-watering, reduced transcript levels of drought stress marker genes, and reduced loss of chlorophyll content in leaves, in combination with stems showing the highest embolism resistance, most pronounced lignification, and thickest intervessel pit membranes. In contrast, the most sensitive Cvi ecotype shows the opposite extreme of the same set of traits. The remaining four genotypes show variations in this drought syndrome. Our results reveal that anatomical, ecophysiological, and molecular adaptations across organs are intertwined, and multiple (differentially combined) strategies can be applied to acquire a certain level of drought tolerance.

miR156-independent repression of the ageing pathway by longevity-promoting AHL proteins in Arabidopsis.

Plants age by developmental phase changes. In Arabidopsis, the juvenile to adult vegetative phase change (VPC) is marked by clear heteroblastic changes in leaves. VPC and the subsequent vegetative to reproductive phase change are promoted by SQUAMOSA PROMOTOR BINDING PROTEIN-LIKE (SPL) transcription factors and repressed by miR156/157 targeting SPL transcripts. By genetic, phenotypic, and gene expression analyses, we studied the role of the longevity-promoting AT-HOOK MOTIF NUCLEAR LOCALIZED 15 (AHL15) and family members in SPL-driven plant ageing. Arabidopsis ahl loss-of-function mutants showed accelerated VPC and flowering, whereas AHL15 overexpression delayed these phase changes. Expression analysis and tissue-specific AHL15 overexpression revealed that AHL15 affects VPC and flowering time directly through its expression in the shoot apical meristem and young leaves, and that AHL15 represses SPL2/9/13/15 gene expression in a miR156/157-independent manner. The juvenile traits of spl loss-of-function mutants appeared to depend on enhanced expression of the AHL15 gene, whereas SPL activity prevented vegetative growth from axillary meristem by repressing AHL15 expression. Our results place AHL15 and close family members together with SPLs in a reciprocal regulatory feedback loop that modulates VPC, flowering time, and axillary meristem development in response to both internal and external signals.

Intervessel pit membrane thickness best explains variation in embolism resistance amongst stems of Arabidopsis thaliana accessions.

BACKGROUND AND AIMS: The ability to avoid drought-induced embolisms in the xylem is one of the essential traits for plants to survive periods of water shortage. Over the past three decades, hydraulic studies have been focusing on trees, which limits our ability to understand how herbs tolerate drought. Here we investigate the embolism resistance in inflorescence stems of four Arabidopsis thaliana accessions that differ in growth form and drought response. We assess functional traits underlying the variation in embolism resistance amongst the accessions studied using detailed anatomical observations. METHODS: Vulnerability to xylem embolism was evaluated via vulnerability curves using the centrifuge technique and linked with detailed anatomical observations in stems using light microscopy and transmission electron microscopy. KEY RESULTS: The data show significant differences in stem P50, varying 2-fold from -1.58 MPa in the Cape Verde Island accession to -3.07 MPa in the woody soc1 ful double mutant. Out of all the anatomical traits measured, intervessel pit membrane thickness (TPM) best explains the differences in P50, as well as P12 and P88. The association between embolism resistance and TPM can be functionally explained by the air-seeding hypothesis. There is no evidence that the correlation between increased woodiness and increased embolism resistance is directly related to functional aspects. However, we found that increased woodiness is strongly linked to other lignification characters, explaining why mechanical stem reinforcement is indirectly related to increased embolism resistance. CONCLUSIONS: The woodier or more lignified accessions are more resistant to embolism than the herbaceous accessions, confirming the link between increased stem lignification and increased embolism resistance, as also observed in other lineages. Intervessel pit membrane thickness and, to a lesser extent, theoretical vessel implosion resistance and vessel wall thickness are the missing functional links between stem lignification and embolism resistance.

Identification of root transcriptional responses to shoot illumination in Arabidopsis thaliana.

KEY MESSAGE: The transcriptional profile of roots is highly affected by shoot illumination. Transcriptogram analysis allows the identification of cellular processes that are not detected by DESeq. Light is a key environmental factor regulating plant growth and development. Arabidopsis thaliana seedlings grown under light display a photomorphogenic development pattern, showing short hypocotyl and long roots. On the other hand, when grown in darkness, they display skotomorphogenic development, with long hypocotyls and short roots. Although many signals from shoots might be important for triggering root growth, the early transcriptional responses that stimulate primary root elongation are still unknown. Here, we aimed to investigate which genes are involved in the early photomorphogenic root development of dark grown roots. We found that 1616 genes 4 days after germination (days-old), and 3920 genes 7 days-old were differently expressed in roots when the shoot was exposed to light. Of these genes, 979 were up regulated in 4 days and 2784 at 7 days-old. We compared the functional categorization of differentially regulated processes by two methods: GO term enrichment and transcriptogram analysis. Expression analysis of nine selected candidate genes in roots confirmed the data observed in the RNA-seq analysis. Loss-of-function mutants of these selected differentially expressed genes suggest the involvement of these genes in root development in response to shoot illumination. Our findings are consistent with the observation that dark grown roots respond to the shoot-perceived aboveground light environment.

Plasticity in Cell Division Patterns and Auxin Transport Dependency during in Vitro Embryogenesis in Brassica napus.

In Arabidopsis thaliana, zygotic embryo divisions are highly regular, but it is not clear how embryo patterning is established in species or culture systems with irregular cell divisions. We investigated this using the Brassica napus microspore embryogenesis system, where the male gametophyte is reprogrammed in vitro to form haploid embryos in the absence of exogenous growth regulators. Microspore embryos are formed via two pathways: a zygotic-like pathway, characterized by initial suspensor formation followed by embryo proper formation from the distal cell of the suspensor, and a pathway characterized by initially unorganized embryos lacking a suspensor. Using embryo fate and auxin markers, we show that the zygotic-like pathway requires polar auxin transport for embryo proper specification from the suspensor, while the suspensorless pathway is polar auxin transport independent and marked by an initial auxin maximum, suggesting early embryo proper establishment in the absence of a basal suspensor. Polarity establishment in this suspensorless pathway was triggered and guided by rupture of the pollen exine. Irregular division patterns did not affect cell fate establishment in either pathway. These results confirm the importance of the suspensor and suspensor-driven auxin transport in patterning, but also uncover a mechanism where cell patterning is less regular and independent of auxin transport.

Modelling the dynamics of polar auxin transport in inflorescence stems of Arabidopsis thaliana.

The polar transport of the plant hormone auxin has been the subject of many studies, several involving mathematical modelling. Unfortunately, most of these models have not been experimentally verified. Here we present experimental measurements of long-distance polar auxin transport (PAT) in segments of inflorescence stems of Arabidopsis thaliana together with a descriptive mathematical model that was developed from these data. It is based on a general advection-diffusion equation for auxin density, as suggested by the chemiosmotic theory, but is extended to incorporate both immobilization of auxin and exchange with the surrounding tissue of cells involved in PAT, in order to account for crucial observations. We found that development of the present model assisted effectively in the analysis of experimental observations. As an example, we discuss the analysis of a quadruple mutant for all four AUX1/LAX1-LAX3 influx carriers genes. We found a drastic change in the parameters governing the exchange of PAT channels with the surrounding tissue, whereas the velocity was still of the order of magnitude of the wild type. In addition, the steady-state flux of auxin through the PAT system of the mutant did not exhibit a saturable component, as we found for the wild type, suggesting that the import carriers are responsible for the saturable component in the wild type. In the accompanying Supplementary data available at JXB online, we describe in more detail the data-driven development of the model, review and derive predictions from a mathematical model of the chemiosmotic theory, and explore relationships between parameters in our model and processes and parameters at the cellular level.

A regulated auxin minimum is required for seed dispersal in Arabidopsis.

Local hormone maxima are essential for the development of multicellular structures and organs. For example, steroid hormones accumulate in specific cell types of the animal fetus to induce sexual differentiation and concentration peaks of the plant hormone auxin direct organ initiation and mediate tissue patterning. Here we provide an example of a regulated local hormone minimum required during organogenesis. Our results demonstrate that formation of a local auxin minimum is necessary for specification of the valve margin separation layer where Arabidopsis fruit opening takes place. Consequently, ectopic production of auxin, specifically in valve margin cells, leads to a complete loss of proper cell fate determination. The valve margin identity factor INDEHISCENT (IND) is responsible for forming the auxin minimum by coordinating auxin efflux in separation-layer cells. We propose that the simplicity of formation and maintenance make local hormone minima particularly well suited to specify a small number of cells such as the stripes at the valve margins.

PIN-driven polar auxin transport in plant developmental plasticity: a key target for environmental and endogenous signals.

Plants master the art of coping with environmental challenges in two ways: on the one hand, through their extensive defense systems, and on the other, by their developmental plasticity. The plant hormone auxin plays an important role in a plant's adaptations to its surroundings, as it specifies organ orientation and positioning by regulating cell growth and division in response to internal and external signals. Important in auxin action is the family of PIN-FORMED (PIN) auxin transport proteins that generate auxin maxima and minima by driving polar cell-to-cell transport of auxin through their asymmetric subcellular distribution. Here, we review how regulatory proteins, the cytoskeleton, and membrane trafficking affect PIN expression and localization. Transcriptional regulation of PIN genes alters protein abundance, provides tissue-specific expression, and enables feedback based on auxin concentrations and crosstalk with other hormones. Post-transcriptional modification, for example by PIN phosphorylation or ubiquitination, provides regulation through protein trafficking and degradation, changing the direction and quantity of the auxin flow. Several plant hormones affect PIN abundance, resulting in another means of crosstalk between auxin and these hormones. In conclusion, PIN proteins are instrumental in directing plant developmental responses to environmental and endogenous signals.

Plasma membrane-bound AGC3 kinases phosphorylate PIN auxin carriers at TPRXS(N/S) motifs to direct apical PIN recycling.

Polar membrane cargo delivery is crucial for establishing cell polarity and for directional transport processes. In plants, polar trafficking mediates the dynamic asymmetric distribution of PIN FORMED (PIN) carriers, which drive polar cell-to-cell transport of the hormone auxin, thereby generating auxin maxima and minima that control development. The Arabidopsis PINOID (PID) protein kinase instructs apical PIN localization by phosphorylating PINs. Here, we identified the PID homologs WAG1 and WAG2 as new PIN polarity regulators. We show that the AGC3 kinases PID, WAG1 and WAG2, and not other plant AGC kinases, instruct recruitment of PINs into the apical recycling pathway by phosphorylating the middle serine in three conserved TPRXS(N/S) motifs within the PIN central hydrophilic loop. Our results put forward a model by which apolarly localized PID, WAG1 and WAG2 phosphorylate PINs at the plasma membrane after default non-polar PIN secretion, and trigger endocytosis-dependent apical PIN recycling. This phosphorylation-triggered apical PIN recycling competes with ARF-GEF GNOM-dependent basal recycling to promote apical PIN localization. In planta, expression domains of PID, WAG1 and WAG2 correlate with apical localization of PINs in those cell types, indicating the importance of these kinases for apical PIN localization. Our data show that by directing polar PIN localization and PIN-mediated polar auxin transport, the three AGC3 kinases redundantly regulate cotyledon development, root meristem size and gravitropic response, indicating their involvement in both programmed and adaptive plant development.

Efficient gene targeting in Penicillium chrysogenum using novel Agrobacterium-mediated transformation approaches.

The industrial production of ß-lactam antibiotics by Penicillium chrysogenum has increased tremendously over the last decades, however, further optimization via classical strain and process improvement has reached its limits. The availability of the genome sequence provides new opportunities for directed strain improvement, but this requires the establishment of an efficient gene targeting (GT) system. Recently, mutations affecting the non-homologous end joining (NHEJ) pathway were shown to increase GT efficiencies following PEG-mediated DNA transfer in P. chrysogenum from 1% to 50%. Apart from direct DNA transfer many fungi can efficiently be transformed using the T-DNA transfer system of the soil bacterium Agrobacterium tumefaciens, however, for P. chrysogenum no robust system for Agrobacterium-mediated transformation was available. We obtained efficient AMT of P. chrysogenum spores with the nourseothricin acetyltransferase gene as selection marker, and using this system we investigated if AMT in a NHEJ mutant background could further enhance GT efficiencies. In general, AMT resulted in higher GT efficiencies than direct DNA transfer, although the final frequencies depended on the Agrobacterium strain and plasmid backbone used. Providing overlapping and complementing fragments on two different plasmid backbones via the same Agrobacterium host was shown to be most effective. This so-called split-marker or bi-partite method resulted in highly efficient GT (>97%) almost exclusively without additional ectopic T-DNA insertions. As this method provides for an efficient GT method independent of protoplasts, it can be applied to other fungi for which no protoplasts can be generated or for which protoplast transformation leads to varying results.

Phosphorylation of conserved PIN motifs directs Arabidopsis PIN1 polarity and auxin transport.

Polar cell-to-cell transport of auxin by plasma membrane-localized PIN-FORMED (PIN) auxin efflux carriers generates auxin gradients that provide positional information for various plant developmental processes. The apical-basal polar localization of the PIN proteins that determines the direction of auxin flow is controlled by reversible phosphorylation of the PIN hydrophilic loop (PINHL). Here, we identified three evolutionarily conserved TPRXS(N/S) motifs within the PIN1HL and proved that the central Ser residues were phosphorylated by the PINOID (PID) kinase. Loss-of-phosphorylation PIN1:green fluorescent protein (GFP) (Ser to Ala) induced inflorescence defects, correlating with their basal localization in the shoot apex, and induced internalization of PIN1:GFP during embryogenesis, leading to strong embryo defects. Conversely, phosphomimic PIN1:GFP (Ser to Glu) showed apical localization in the shoot apex but did not rescue pin1 inflorescence defects. Both loss-of-phosphorylation and phosphomimic PIN1:GFP proteins were insensitive to PID overexpression. The basal localization of loss-of-phosphorylation PIN1:GFP increased auxin accumulation in the root tips, partially rescuing PID overexpression-induced root collapse. Collectively, our data indicate that reversible phosphorylation of the conserved Ser residues in the PIN1HL by PID (and possibly by other AGC kinases) is required and sufficient for proper PIN1 localization and is thus essential for generating the differential auxin distribution that directs plant development.

Physiological and morphological changes during early and later stages of fruit growth in Capsicum annuum.

Fruit-set involves a series of physiological and morphological changes that are well described for tomato and Arabidopsis, but largely unknown for sweet pepper (Capsicum annuum). The aim of this paper is to investigate whether mechanisms of fruit-set observed in Arabidopsis and tomato are also applicable to C. annuum. To do this, we accurately timed the physiological and morphological changes in a post-pollinated and un-pollinated ovary. A vascular connection between ovule and replum was observed in fertilized ovaries that undergo fruit development, and this connection was absent in unfertilized ovaries that abort. This indicates that vascular connection between ovule and replum is an early indicator for successful fruit development after pollination and fertilization. Evaluation of histological changes in the carpel of a fertilized and unfertilized ovary indicated that increase in cell number and cell diameter both contribute to early fruit growth. Cell division contributes more during early fruit growth while cell expansion contributes more at later stages of fruit growth in C. annuum. The simultaneous occurrence of a peak in auxin concentration and a strong increase in cell diameter in the carpel of seeded fruits suggest that indole-3-acetic acid stimulates a major increase in cell diameter at later stages of fruit growth. The series of physiological and morphological events observed during fruit-set in C. annuum are similar to what has been reported for tomato and Arabidopsis. This indicates that tomato and Arabidopsis are suitable model plants to understand details of fruit-set mechanisms in C. annuum.

Phosphorylation-dependent trafficking of plasma membrane proteins in animal and plant cells.

In both unicellular and multicellular organisms, transmembrane (TM) proteins are sorted to and retained at specific membrane domains by endomembrane trafficking mechanisms that recognize sorting signals in the these proteins. The trafficking and distribution of plasma membrane (PM)-localized TM proteins (PM proteins), especially of those PM proteins that show an asymmetric distribution over the PM, has received much attention, as their proper PM localization is crucial for elementary signaling and transport processes, and defects in their localization often lead to severe disease symptoms or developmental defects. The subcellular localization of PM proteins is dynamically regulated by post-translational modifications, such as phosphorylation and ubiquitination. These modificaitons mostly occur on sorting signals that are located in the larger cytosolic domains of the cargo proteins. Here we review the effects of phosphorylation of PM proteins on their trafficking, and present the key examples from the animal field that have been subject to studies for already several decades, such as that of aquaporin 2 and the epidermal growth factor receptor. Our knowledge on cargo trafficking in plants is largely based on studies of the family of PIN FORMED (PIN) carriers that mediate the efflux of the plant hormone auxin. We will review what is known on the subcellular distribution and trafficking of PIN proteins, with a focus on how this is modulated by phosphorylation, and identify and discuss analogies and differences in trafficking with the well-studied animal examples.

Polar auxin transport: an early invention.

In higher plants, cell-to-cell polar auxin transport (PAT) of the phytohormone auxin, indole-3-acetic acid (IAA), generates maxima and minima that direct growth and development. Although IAA is present in all plant phyla, PAT has only been detected in land plants, the earliest being the Bryophytes. Charophyta, a group of freshwater green algae, are among the first multicellular algae with a land plant-like phenotype and are ancestors to land plants. IAA has been detected in members of Charophyta, but its developmental role and the occurrence of PAT are unknown. We show that naphthylphthalamic acid (NPA)-sensitive PAT occurs in internodal cells of Chara corallina. The relatively high velocity (at least 4-5 cm/h) of auxin transport through the giant (3-5 cm) Chara cells does not occur by simple diffusion and is not sensitive to a specific cytoplasmic streaming inhibitor. The results demonstrate that PAT evolved early in multicellular plant life. The giant Chara cells provide a unique new model system to study PAT, as Chara allows the combining of real-time measurements and mathematical modelling with molecular, developmental, cellular, and electrophysiological studies.