The effector AvrRxo1 phosphorylates NAD in planta

Abstract

Gram-negative bacterial pathogens of plants and animals employ type III secreted effectors to suppress innate immunity. Most characterized effectors work through modification of host proteins or transcriptional regulators, although a few are known to modify small molecule targets. The Xanthomonas type III secreted avirulence factor AvrRxo1 is a structural homolog of the zeta toxin family of sugar-nucleotide kinases that suppresses bacterial growth. AvrRxo1 was recently reported to phosphorylate the central metabolite and signaling molecule NAD in vitro, suggesting that the effector might enhance bacterial virulence on plants through manipulation of primary metabolic pathways. In this study, we determine that AvrRxo1 phosphorylates NAD in planta, and that its kinase catalytic sites are necessary for its toxic and resistance-triggering phenotypes. A global metabolomics approach was used to independently identify 3’-NADP as the sole detectable product of AvrRxo1 expression in yeast and bacteria, and NAD kinase activity was confirmed in vitro. 3’-NADP accumulated upon transient expression of AvrRxo1 in Nicotiana benthamiana and in rice leaves infected with avrRxo1-expressing strains of X. oryzae. Mutation of the catalytic aspartic acid residue D193 abolished AvrRxo1 kinase activity and several phenotypes of AvrRxo1, including toxicity in yeast, bacteria, and plants, suppression of the flg22-triggered ROS burst, and ability to trigger an R gene-mediated hypersensitive response. A mutation in the Walker A ATP-binding motif abolished the toxicity of AvrRxo1, but did not abolish the 3’-NADP production, virulence enhancement, ROS suppression, or HR-triggering phenotypes of AvrRxo1. These results demonstrate that a type III effector targets the central metabolite and redox carrier NAD in planta, and that this catalytic activity is required for toxicity and suppression of the ROS burst.

Differential localization of Hessian fly candidate effectors in resistant and susceptible wheat plants

“…One of the difficulties for studying insect effectors is that it is hard to detect them in host tissues due to the small amounts of insect proteins in host plants (Wang et al, 2018). So far only a handful of effector proteins from insect and pathogen species have been directly visualized in infested plants, including the pea aphid effector Armet in infested fava bean (Wang et al, 2015), aphid effectors Mp10, MpOS-D1, MpC002, and MpPIntO1 in infested Arabidopsis thaliana (Mugford et al, 2016), an endo-β-1,4-glucanase and an EFhand calcium-binding protein from the brown planthopper in infested rice plants (Ye et al, 2017;Ji et al, 2017), the low molecular weight whitefly salivary protein Bt56 in tobacco plants (Xu et al, 2019), and the effector AvrRxo1 in rice tissues from Xanthomonas oryzae (Shidore et al 2017).…”

Convergent Evolution of Pathogen Effectors toward Reactive Oxygen Species Signaling Networks in Plants

“…The X. oryzae pv. oryzicola effector AvrRxo1 targets and phosphorylates the central metabolite and redox carrier NAD in planta , and this catalytic activity is required for suppression of the ROS burst ( Shidore et al, 2017 ). The X. campestris pv.…”

Roles of Nicotinamide Adenine Dinucleotide (NAD+) in Biological Systems

“…In support of the importance of NAD + in plant immunity, it was shown that the effector AvrRxo1 phosphorylates NAD deactivating the immune response in plants [90].…”