Abstract
Many plant bacterial pathogens including Pseudomonas species, utilize the type III secretion system (T3SS) to deliver effector proteins into plant cells. Genes encoding the T3SS and its effectors are repressed in nutrient-rich media but are rapidly induced after the bacteria enter a plant or are transferred into nutrient-deficient media. To understand how the T3SS genes are regulated, we screened for P. savastanoi pv. phaseolicola (Psph) mutants displaying diminished induction of avrPto-luc, a reporter for the T3SS genes, in Arabidopsis. A mutant carrying transposon insertion into a gene coding for a small functional unknown protein, designated as rhpC, was identified that poorly induced avrPto-luc in plants and in minimal medium (MM). Interestingly, rhpC is located immediately downstream of a putative metalloprotease gene named rhpP, and the two genes are organized in an operon rhpPC; but rhpP and rhpC displayed different RNA expression patterns in nutrient-rich King’s B medium (KB) and MM. Deletion of the whole rhpPC locus did not significantly affect the avrPto-luc induction, implying coordinated actions of rhpP and rhpC in regulating the T3SS genes. Further analysis showed that RhpC was a cytoplasmic protein that interacted with RhpP and targeted RhpP to the periplasm. In the absence of RhpC, RhpP was localized in the cytoplasm and caused a reduction of HrpL, a key regulator of the T3SS genes, and also reduced the fitness of Psph. Expression of RhpP alone in E. coli inhibited the bacterial growth. The detrimental effect of RhpP on the fitness of Psph and E. coli required metalloprotease active sites, and was repressed when RhpC was co-expressed with RhpP. The coordination between rhpP and rhpC in tuning the T3SS gene expression and cell fitness reveals a novel regulatory mechanism for bacterial pathogenesis. The function of RhpP in the periplasm remains to be studied.
Highlights
Many Gram-negative bacterial pathogens rely on the T3SS for successful infection of their hosts [1]
We discovered that the bicistronic genes in the rhpPC locus of Psph act coordinately to regulate the T3SS gene expression, bacterial fitness, and pathogenicity. rhpP encodes a metalloprotease that can degrade the key T3SS regulator protein HrpL and reduce bacterial fitness. rhpC encodes a chaperone that inhibits the RhpP activity and mediates translocation of RhpP to the periplasm
The level of rhpP RNA is high in King’s B medium (KB) but decreases in MM, but the rhpC RNA is low in KB but increases in MM
Summary
Many Gram-negative bacterial pathogens rely on the T3SS for successful infection of their hosts [1]. The T3SS is encoded by a cluster of hrp/hrc genes that are essential for the induction of a hypersensitive response (HR) in resistant and nonhost plants and pathogenicity in susceptible plants [2]. The T3SS functions as a conduit to deliver an array of effector proteins into plant cells [3]. The expression of hrp/hrc genes and the effector genes (together called T3SS genes hereafter) is coordinately regulated by various environmental and host factors [6]. T3SS genes are expressed at a very low level when grown in nutrient-rich medium, but quickly induced to high levels after the bacteria are infiltrated into plants or cultured in nutrient-deficient medium that is believed to resemble the environment of the plant intercellular spaces where the bacteria proliferate during infection [7, 8]
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