Systematic Y2H Screening Reveals Extensive Effector-Complex Formation

Abstract

During infection pathogens secrete small molecules, termed effectors, to manipulate and control the interaction with their specific hosts. Both the pathogen and the plant are under high selective pressure to rapidly adapt and co-evolve in what is usually referred to as molecular arms race. Components of the host’s immune system form a network that processes information about molecules with a foreign origin and damage-associated signals, integrating them with developmental and abiotic cues to adapt the plant’s responses. Both in the case of nucleotide-binding leucine-rich repeat receptors and leucine-rich repeat receptor kinases interaction networks have been extensively characterized. However, little is known on whether pathogenic effectors form complexes to overcome plant immunity and promote disease. Ustilago maydis, a biotrophic fungal pathogen that infects maize plants, produces effectors that target hubs in the immune network of the host cell. Here we assess the capability of U. maydis effector candidates to interact with each other, which may play a crucial role during the infection process. Using a systematic yeast-two-hybrid approach and based on a preliminary pooled screen, we selected 63 putative effectors for one-on-one matings with a library of nearly 300 effector candidates. We found that 126 of these effector candidates interacted either with themselves or other predicted effectors. Although the functional relevance of the observed interactions remains elusive, we propose that the observed abundance in complex formation between effectors adds an additional level of complexity to effector research and should be taken into consideration when studying effector evolution and function. Based on this fundamental finding, we suggest various scenarios which could evolutionarily drive the formation and stabilization of an effector interactome.

TOPLESS promotes plant immunity by repressing auxin signaling and is targeted by the fungal effector Naked1

“…Besides a second unidentified host target, another explanation for the virulence function of Nkd1 could be its interaction with one or more additional effectors which depend on Nkd1 to perform their function in maize. Systematic Y2H screens between U. maydis effectors indicate a massive effector interactome (Alcantara et al, 2019) and effector-effector interactions have been shown to impact virulence in other pathosystems (Tzfira et al, 2004, Magori and Citovsky, 2011, Cain et al, 2008. Even though we could not detect an interaction between Nkd1 and Jsi1 (Alcantara et al, 2019), Nkd1 could potentially interact with other U. maydis effectors during maize infection.…”

“…Systematic Y2H screens between U. maydis effectors indicate a massive effector interactome (Alcantara et al, 2019) and effector-effector interactions have been shown to impact virulence in other pathosystems (Tzfira et al, 2004, Magori and Citovsky, 2011, Cain et al, 2008. Even though we could not detect an interaction between Nkd1 and Jsi1 (Alcantara et al, 2019), Nkd1 could potentially interact with other U. maydis effectors during maize infection. Therefore, analysis of Nkd1 interaction partners (plant or fungal) by IP-MS from infected maize tissue could help identify its second, non-redundant function.…”

The Sporisorium reilianum Effector Vag2 Promotes Head Smut Disease via Suppression of Plant Defense Responses

“…The complex is formed during biotrophic growth and may be involved in effector translocation into plant cells [ 65 ]. A systematic yeast two-hybrid interaction screen of U. maydis effectors revealed extensive complex formation capacity, both by homo- as well as heterodimerization [ 66 ].…”

“…This indicates that each of the three genes individually affects virulence and that their role in virulence is independent of the presence of the other two genes. Of the five U. maydis tin1 genes, two were positive in the yeast two-hybrid effector interaction screen, and for one of them, tin1-2 , three putative interaction partners were retrieved [ 66 ]. Since these are identified as unrelated proteins, there is also no indication of complex formation between different Tin1 proteins in U. maydis .…”

A LysM Domain-Containing Protein LtLysM1 Is Important for Vegetative Growth and Pathogenesis in Woody Plant Pathogen Lasiodiplodia theobromae

“…Another two LysM receptor molecules OsCERK1 with CEBiP were able to form a hetero-oligomer complex to induce downstream immune responses ( Shimizu et al, 2010 ). Plant pathogenic effectors form complexes to overcome plant immunity to promote disease by interacting with themselves or with other effector proteins ( Alcântara et al, 2019 ). PsCRN63 in P. sojae was capable of suppressing PCD associated with PTI via self-dimerization ( Li et al, 2016 ).…”