The Xanthomonas campestris Type III Effector XopJ Proteolytically Degrades Proteasome Subunit RPT6

Abstract

Many animal and plant pathogenic bacteria inject type III effector (T3E) proteins into their eukaryotic host cells to suppress immunity. The Yersinia outer protein J (YopJ) family of T3Es is a widely distributed family of effector proteins found in both animal and plant pathogens, and its members are highly diversified in virulence functions. Some members have been shown to possess acetyltransferase activity; however, whether this is a general feature of YopJ family T3Es is currently unknown. The T3E Xanthomonas outer protein J (XopJ), a YopJ family effector from the plant pathogen Xanthomonas campestris pv vesicatoria, interacts with the proteasomal subunit Regulatory Particle AAA-ATPase6 (RPT6) in planta to suppress proteasome activity, resulting in the inhibition of salicylic acid-related immune responses. Here, we show that XopJ has protease activity to specifically degrade RPT6, leading to reduced proteasome activity in the cytoplasm as well as in the nucleus. Proteolytic degradation of RPT6 was dependent on the localization of XopJ to the plasma membrane as well as on its catalytic triad. Mutation of the Walker B motif of RPT6 prevented XopJ-mediated degradation of the protein but not XopJ interaction. This indicates that the interaction of RPT6 with XopJ is dependent on the ATP-binding activity of RPT6, but proteolytic cleavage additionally requires its ATPase activity. Inhibition of the proteasome impairs the proteasomal turnover of Nonexpressor of Pathogenesis-Related1 (NPR1), the master regulator of salicylic acid responses, leading to the accumulation of ubiquitinated NPR1, which likely interferes with the full induction of NPR1 target genes. Our results show that YopJ family T3Es are not only highly diversified in virulence function but also appear to possess different biochemical activities.

The Proteasome Acts as a Hub for Plant Immunity and Is Targeted by Pseudomonas Type III Effectors

“…However, XopJ, a T3E from X. campestris pv vesicatoria 85-10, was shown to proteolytically degrade the 19S RP subunit RPT6 in order to inhibit proteasome activity and to interfere with SA-mediated defense in susceptible pepper host plants (Üstün et al, 2013;Üstün and Börnke, 2015). A similar mechanism has been proposed for the T3E HopZ4 from P. syringae pv lachrymans (Üstün et al, 2014).…”

“…The first bacterial T3Es identified that directly target the proteasome for defense suppression are XopJ from Xanthomonas campestris pv vesicatoria and HopZ4 from Pseudomonas syringae pv lachrymans. Both closely related T3Es interact with the proteasomal component RPT6, a subunit of the 19S RP, to inhibit proteasome activity (Üstün et al, 2013Üstün and Börnke, 2015). In effect, this results in impaired turnover of the SA master regulator NPR1 and the attenuation of SAdependent defense responses.…”

The type III effector AvrXccB in Xanthomonas campestris pv. campestris targets putative methyltransferases and suppresses innate immunity in Arabidopsis

“…Xanthomonas XopJ and HopZ4, a YopJ family protein from P. syringae , inhibit proteasome activity through interaction with the proteasomal subunit RPT6. The inhibitory effect is dependent on the plasma membrane localization and conserved catalytic triad of these effector proteins (Üstün and Börnke, ; Üstün et al ., ). Therefore, we investigated the effect of the N ‐myristoylation motif on various facets of AvrXccB function.…”

“…ACIP1 acetylation in planta alters its defence function (Cheong et al ., ). XopJ targets the host cell proteasome to suppress salicylic acid‐mediated plant defence (Üstün and Börnke, ; Üstün et al ., ). AvrXv4, like XopD, possesses a SUMO isopeptidase activity, and ectopic expression of AvrXv4 in plants results in a reduction in SUMO‐modified proteins (Roden et al ., ).…”

Behind the lines–actions of bacterial type III effector proteins in plant cells

“…vesicatoria xopJ deletion mutant led to increased proteasome activity when compared with leaves infected with the wild-type strain (Üstün, Bartetzko and Börnke 2013). XopJ interacts with and degrades the ATPase RPT6 (regulatory particle ATPase 6) of the 19S regulatory particle of the 26S proteasome at the plant plasma membrane (Üstün, Bartetzko and Börnke 2013; Üstün and Börnke 2015) (Fig. 4F).…”