Vascularization and nutrient delivery at root-knot nematode feeding sites in host roots

Plants are constantly challenged by pathogens and pests, which can have a profound impact on the yield and quality of produce in agricultural systems. The vascular system of higher plants is critical for growth and for their ability to counteract changing external conditions, serving as a distribution network for water, nutrients, and photosynthates from the source organs to regions where they are in demand. Unfortunately, these features also make it an attractive target for pathogens and pests that demand access to a reliable supply of host resources. The vascular tissue of plants therefore often plays a central role in pathogen and parasite interactions. One of the more striking rearrangements of the host vascular system occurs during root-knot nematode infestation of plant roots. These sedentary endoparasites induce permanent feeding sites that are comprised of ‘giant cells’ and are subject to extensive changes in vascularization, resulting in the giant cells being encaged within a network of de novo formed xylem and phloem cells. Despite being considered critical to the function of the feeding site, the mechanisms underlying this vascularization have received surprisingly little attention when compared with the amount of research on giant cell development and function. An overview of the current knowledge on vascularization of root-knot nematode feeding sites is provided here and recent advances in our understanding of the transport mechanisms involved in nutrient delivery to these parasite-induced sinks are described

Activation and Polarity Control of PIN-FORMED Auxin Transporters by Phosphorylation.

Auxin controls almost every aspect of plant development. Auxin is distributed within the plant by passive diffusion and active cell-to-cell transport. PIN-FORMED (PIN) auxin efflux transporters are polarly distributed in the plasma membranes of many cells, and knowledge about their distribution can predict auxin transport and explain auxin distribution patterns, even in complex tissues. Recent studies have revealed that phosphorylation is essential for PIN activation, suggesting that PIN phosphorylation needs to be taken into account in understanding auxin transport. These findings also ask for a re-examination of previously proposed mechanisms for phosphorylation-dependent PIN polarity control. We provide a comprehensive summary of the current knowledge on PIN regulation by phosphorylation, and discuss possible mechanisms of PIN polarity control in the context of recent findings

A molecular rheostat adjusts auxin flux to promote root protophloem differentiation.

Auxin influences plant development through several distinct concentration-dependent effects <sup>1</sup> . In the Arabidopsis root tip, polar auxin transport by PIN-FORMED (PIN) proteins creates a local auxin accumulation that is required for the maintenance of the stem-cell niche <sup>2-4</sup> . Proximally, stem-cell daughter cells divide repeatedly before they eventually differentiate. This developmental gradient is accompanied by a gradual decrease in auxin levels as cells divide, and subsequently by a gradual increase as the cells differentiate <sup>5,6</sup> . However, the timing of differentiation is not uniform across cell files. For instance, developing protophloem sieve elements (PPSEs) differentiate as neighbouring cells still divide. Here we show that PPSE differentiation involves local steepening of the post-meristematic auxin gradient. BREVIS RADIX (BRX) and PROTEIN KINASE ASSOCIATED WITH BRX (PAX) are interacting plasma-membrane-associated, polarly localized proteins that co-localize with PIN proteins at the rootward end of developing PPSEs. Both brx and pax mutants display impaired PPSE differentiation. Similar to other AGC-family kinases, PAX activates PIN-mediated auxin efflux, whereas BRX strongly dampens this stimulation. Efficient BRX plasma-membrane localization depends on PAX, but auxin negatively regulates BRX plasma-membrane association and promotes PAX activity. Thus, our data support a model in which BRX and PAX are elements of a molecular rheostat that modulates auxin flux through developing PPSEs, thereby timing PPSE differentiation

Mapping and engineering of auxin-induced plasma membrane dissociation in BRX family proteins.

Angiosperms have evolved the phloem for the long-distance transport of metabolites. The complex process of phloem development involves genes that only occur in vascular plant lineages. For example, in Arabidopsis thaliana, the BREVIS RADIX (BRX) gene is required for continuous root protophloem differentiation, together with PROTEIN KINASE ASSOCIATED WITH BRX (PAX). BRX and its BRX-LIKE (BRXL) homologs are composed of four highly conserved domains including the signature tandem BRX domains that are separated by variable spacers. Nevertheless, BRX family proteins have functionally diverged. For instance, BRXL2 can only partially replace BRX in the root protophloem. This divergence is reflected in physiologically relevant differences in protein behavior, such as auxin-induced plasma membrane dissociation of BRX, which is not observed for BRXL2. Here we dissected the differential functions of BRX family proteins using a set of amino acid substitutions and domain swaps. Our data suggest that the plasma membrane-associated tandem BRX domains are both necessary and sufficient to convey the biological outputs of BRX function and therefore constitute an important regulatory entity. Moreover, PAX target phosphosites in the linker between the two BRX domains mediate the auxin-induced plasma membrane dissociation. Engineering these sites into BRXL2 renders this modified protein auxin-responsive and thereby increases its biological activity in the root protophloem context

Plasma Membrane Domain Patterning and Self-Reinforcing Polarity in Arabidopsis.

Cell polarity is a key feature in the development of multicellular organisms. For instance, asymmetrically localized plasma-membrane-integral PIN-FORMED (PIN) proteins direct transcellular fluxes of the phytohormone auxin that govern plant development. Fine-tuned auxin flux is important for root protophloem sieve element differentiation and requires the interacting plasma-membrane-associated BREVIS RADIX (BRX) and PROTEIN KINASE ASSOCIATED WITH BRX (PAX) proteins. We observed "donut-like" polar PIN localization in developing sieve elements that depends on complementary, "muffin-like" polar localization of BRX and PAX. Plasma membrane association and polarity of PAX, and indirectly BRX, largely depends on phosphatidylinositol-4,5-bisphosphate. Consistently, mutants in phosphatidylinositol-4-phosphate 5-kinases (PIP5Ks) display protophloem differentiation defects similar to brx mutants. The same PIP5Ks are in complex with BRX and display "muffin-like" polar localization. Our data suggest that the BRX-PAX module recruits PIP5Ks to reinforce PAX polarity and thereby the polarity of all three proteins, which is required to maintain a local PIN minimum