Partner control in arbuscular mycorrhizal symbiosis

The arbuscular mycorrhizal (AM) symbiosis is the most widespread symbiotic association between land plants and arbuscular mycorrhizal fungi belonging to Glomeromycotina subphylum. This mutualistic association can have a range of benefits for the plant, such as increased access to mineral nutrients (especially phosphate and nitrogen) and water in the soil as well as protection from various biotic and abiotic stresses. In return for these services, the fungi receive fixed carbon from their hosts to complete their life cycle. To keep this interaction beneficial, multiple layers of control are exerted by both partners at different stages of the interaction. In this thesis, we studied the role of AM fungal effectors to facilitate colonization and how host plants sense how much phosphate they receive from their partner to control the development of the arbuscules.Similar to pathogenic fungi, the cell walls of AM fungi contain common microbe-associated molecular patterns (MAMPs), such as chitin and β-glucans, that can trigger plant immune responses. The extreme host range of AM fungi raises the question how they can efficiently deal with the immune system of such a wide variety of plants. It has recently become clear that secreted AM fungal effector proteins can play a key role. In chapter 2, we studied the role of a secreted nuclear-localization signal containing effector from Rhizophagus irregularis, called Nuclear Localized Effector1 (RiNLE1), which is highly and specifically expressed in arbuscules in wide variety of plant species. RiNLE1 was able translocate to the host nucleus where it could interact with the core nucleosome component Histone 2B (H2B) and impair its mono-ubiquitination. This resulted in the suppression of defense-related gene expression and enhanced colonization levels. This chapter highlighted a novel mechanism by which AM fungi can effectively affect the plants epigenome via direct interaction with a core nucleosome component to facilitate colonization. RiNLE1-like effectors were found in a range of fungi that establish intimate interactions with plants, suggesting that this type of effector may be more widely recruited to manipulate host defense responses.Despite the fact that there seems to be little to no strict host specificity in the AM interaction. so-called host preferences have been widely observed in the field. In chapter 3, we studied an effector from Rhizophagus irregularis DAOM197198, called RiFGB1, that is expressed in a host-dependent manner. RiFGB1 is highly induced in the interaction with Allium schoenoprasum (chives) as host plant but not in the interaction with dicot hosts like Medicago truncatula and Nicotiana benthamiana. RiFGB1 was shown to be a homolog of the Piriformospora indica fungal β-glucan-binding lectin (FGB1). We found that RiFGB1 can bind a variety of glycan molecules such as β-glucan, chitin and xylan. Furthermore RiFGB1 could interfere with a wide range of MAMP-triggered immune responses. Furthermore, different expression levels of FGB1 homologs in another R. irregularis isolate seemed to correlate with a reduced colonization of this isolate on chives. These data suggest that effectors may contribute to host preferences through their ability to regulate the host immune system.To keep the symbiosis beneficial a plant must be able to sense how much nutrients it obtains from its partner and to integrate it with its needs. For example, if the fungus fails to provide phosphate the plant can terminate the interaction by degrading the arbuscules to prevent a carbon drain by the fungus. How plants sense the phosphate status in arbuscule-containing cells and accordingly control arbuscule lifetime or function is therefore a major question. In chapter 4, we identified two Medicago truncatula phosphate-sensing SPX-domain containing proteins, SPX1 and SPX3, as key regulators of phosphate starvation responses as well as fungal colonization and arbuscule degradation. SPX1 and SPX3 are induced by phosphate starvation but become restricted to arbuscule-containing cells upon establishment of AM symbiosis. Under non-symbiotic conditions they control phosphate homeostasis in part through their interaction with the phosphate starvation response factor PHR2. Furthermore, upon phosphate deficiency they facilitate expression of the strigolactone biosynthesis gene DWARF27, which correlates with increased fungal branching by root exudates and increased root colonization. Later, in the arbuscule-containing cells SPX1 and SPX3 redundantly control the timely degradation of arbuscules.To understand how SPX1 and SPX3 control arbuscule lifetime, we searched for interacting partners in chapter 5. We did not find any interaction with the known transcriptional regulators of arbuscule degradation MYB1, NSP1 and DELLA. Instead, through co-immunoprecipitation coupled with liquid chromatography–mass spectrometry, we identified PIP2 aquaporins as interactors of both SPX1 and SPX3. Overexpression of MtPIP2;7 reduced overall arbuscule abundance but also caused a higher “good” to “degrading” arbuscule ratio, suggesting reduced arbuscule degradation. These results now offer a basis to study the intriguing possibility that SPX1 and SPX3 help to regulate phosphate transport and/or arbuscule degeneration through the regulation of PIP2 activity.In chapter 6, the results generated during my thesis research are discussed in the broader perspective of partner preferences during AM symbiosis. It has become clear that complex and interconnected signaling pathways concerning nutrient status and defense responses are integrated to control AM symbiosis development. Extending this knowledge will be important realize to the widely recognized potential of AM fungi as sustainable biofertilizers.