The growth-promoting and root-colonizing endophyte Piriformospora indica induces camalexin and the expression of CYP79B2, CYP79B3, CYP71A13, PAD3, and WRKY33 required for the synthesis of indole-3-acetaldoxime (IAOx)-derived compounds in the roots of Arabidopsis seedlings. Upregulation of the mRNA levels by P. indica requires cytoplasmic calcium elevation and mitogen-activated protein kinase 3 but not root-hair-deficient 2, radical oxygen production, or the 3-phosphoinositide-dependent kinase 1/oxidative signal-inducible 1 pathway. Because P. indica-mediated growth promotion is impaired in cyp79B2 cyp79B3 seedlings, while pad3 seedlings-which do not accumulate camalexin-still respond to the fungus, IAOx-derived compounds other than camalexin (e.g., indole glucosinolates) are required during early phases of the beneficial interaction. The roots of cyp79B2 cyp79B3 seedlings are more colonized than wild-type roots, and upregulation of the defense genes pathogenesis-related (PR)-1, PR-3, PDF1.2, phenylalanine ammonia lyase, and germin indicates that the mutant responds to the lack of IAOx-derived compounds by activating other defense processes. After 6 weeks on soil, defense genes are no longer upregulated in wild-type, cyp79B2 cyp79B3, and pad3 roots. This results in uncontrolled fungal growth in the mutant roots and reduced performance of the mutants. We propose that a long-term harmony between the two symbionts requires restriction of root colonization by IAOx-derived compounds.

The vast majority of plants live in symbiotic interaction with mycorrhizal fungi. The fungus delivers soil nutrients to the plant (Javot et al. 2007a, b;Bucher et al. 2009;Adesemoye and Kloepper 2009) while the plant is responsible for the photoassimilates to the fungus (Nehls 2008;Nehls et al. 2010). Mycorrhizal interactions are difficult to investigate at the molecular level because most of the classical model plants do not form mycorrhizal association. As an alternative, endophytic interaction of axenically cultivable Piriformospora indica with the model plant Arabidopsis thaliana might help to understand the basis of the beneficial interactions between two symbionts.We focus on the identification of Arabidopsis mutants which are impaired in establishing or maintaining the interaction in a beneficial mode. For those studies, reproducible and quantitative cocultivation parameters are required. Therefore, we have developed standardized (co)cultivation conditions for the two symbionts, which are described here in details. The mode of interaction and the reproducibility of the data are highly dependent on the quality of the biological material and the cocultivation conditions. Most important parameters are the age and the density of the fungal mycelium, the age of the seedlings and plants, the ratio between the two symbiotic partners, and the cocultivation conditions like temperature, humidity, light intensity, spectral distribution, and photoperiod (Shahollari et al.

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