Genetic evidence for auxin involvement in arbuscular mycorrhiza initiation.

• Formation of arbuscular mycorrhiza (AM) is controlled by a host of small, diffusible signaling molecules, including phytohormones. To test the hypothesis that the plant hormone auxin controls mycorrhiza development, we assessed mycorrhiza formation in two mutants of tomato (Solanum lycopersicum): diageotropica (dgt), an auxin-resistant mutant, and polycotyledon (pct), a mutant with hyperactive polar auxin transport. • Mutant and wild-type (WT) roots were inoculated with spores of the AM fungus Glomus intraradices. Presymbiotic root-fungus interactions were observed in root organ culture (ROC) and internal fungal colonization was quantified both in ROC and in intact seedlings. • In ROC, G. intraradices stimulated presymbiotic root branching in pct but not in dgt roots. pct roots stimulated production of hyphal fans indicative of appressorium formation and were colonized more rapidly than WT roots. By contrast, approaching hyphae reversed direction to grow away from cultured dgt roots and failed to colonize them. In intact seedlings, pct and dgt roots were colonized poorly, but development of hyphae, arbuscules, and vesicles was morphologically normal within roots of both mutants. • We conclude that auxin signaling within host roots is required for the early stages of AM formation, including during presymbiotic signal exchange.

2,4-diacetylphloroglucinol alters plant root development.

Pseudomonas fluorescens isolates containing the phlD gene can protect crops from root pathogens, at least in part through production of the antibiotic 2,4-diacetylphloroglucinol (DAPG). However, the action mechanisms of DAPG are not fully understood, and effects of this antibiotic on host root systems have not been characterized in detail. DAPG inhibited primary root growth and stimulated lateral root production in tomato seedlings. Roots of the auxin-resistant diageotropica mutant of tomato demonstrated reduced DAPG sensitivity with regards to inhibition of primary root growth and induction of root branching. Additionally, applications of exogenous DAPG, at concentrations previously found in the rhizosphere of plants inoculated with DAPG-producing pseudomonads, inhibited the activation of an auxin-inducible GH3 promoter::luciferase reporter gene construct in transgenic tobacco hypocotyls. In this model system, supernatants of 17 phlD+ P. fluorescens isolates had inhibitory effects on luciferase activity similar to synthetic DAPG. In addition, a phlD() mutant strain, unable to produce DAPG, demonstrated delayed inhibitory effects compared with the parent wild-type strain. These results indicate that DAPG can alter crop root architecture by interacting with an auxin-dependent signaling pathway.

Cytokinin inhibits a subset of diageotropica-dependent primary auxin responses in tomato.

Many aspects of plant development are regulated by antagonistic interactions between the plant hormones auxin and cytokinin, but the molecular mechanisms of this interaction are not understood. To test whether cytokinin controls plant development through inhibiting an early step in the auxin response pathway, we compared the effects of cytokinin with those of the dgt (diageotropica) mutation, which is known to block rapid auxin reactions of tomato (Lycopersicon esculentum) hypocotyls. Long-term cytokinin treatment of wild-type seedlings phenocopied morphological traits of dgt plants such as stunting of root and shoot growth, reduced elongation of internodes, reduced apical dominance, and reduced leaf size and complexity. Cytokinin treatment also inhibited rapid auxin responses in hypocotyl segments: auxin-stimulated elongation, H(+) secretion, and ethylene synthesis were all inhibited by cytokinin in wild-type hypocotyl segments, and thus mimicked the impaired auxin responsiveness found in dgt hypocotyls. However, cytokinin failed to inhibit auxin-induced LeSAUR gene expression, an auxin response that is affected by the dgt mutation. In addition, cytokinin treatment inhibited the auxin induction of only one of two 1-aminocyclopropane-1-carboxylic acid synthase genes that exhibited impaired auxin inducibility in dgt hypocotyls. Thus, cytokinin inhibited a subset of the auxin responses impaired in dgt hypocotyls, suggesting that cytokinin blocks at least one branch of the DGT-dependent auxin response pathway.

Developmental regulation of H+-ATPase-dependent auxin responses in the diageotropica mutant of tomato (Lycopersicon esculentum).

Rapid auxin effects on H+ pumping across the plasma membrane precede auxin-induced elongation growth of hypocotyls and swelling of guard cells, as well as auxin inhibition of root growth. To investigate whether auxin-signalling mechanisms in such diverse cell types are similar, we characterized these responses in various tissues of the diageotropica (dgt) mutant of tomato (Lycopersicon esculentum Mill.). Abraded hypocotyl segments of 4-day-old, etiolated dgt seedlings showed an impaired H+ secretion response to applied auxin. mRNA levels for two PM H+-ATPase isoforms, LHA2 and LHA4, were not reduced in dgt hypocotyl segments as compared to wild-type segments, suggesting that the dgt mutation does not affect H+ secretion by reducing the transcription of major PM H+-ATPase genes. The dgt mutation also disrupted auxin inhibition of growth and H+ secretion in roots of 4-day-old dgt seedlings. However, immediately after germination, dgt seedling roots responded to auxin with a near-normal inhibition of growth. In addition, stomata in epidermal peels from 2-week-old dgt cotyledons demonstrated normal auxin-induced opening. We conclude that an intact DGT gene product is required for auxin-induced H+ secretion in tomato hypocotyl segments and for auxin inhibition of H+ secretion in roots of older seedlings, but that a DGT-independent pathway for auxin responses exists in young root tips and in guard cells. A developmentally controlled switch from DGT-independent to DGT-dependent auxin signalling appears to take place in root tips within 2 days after germination.