Abstract
The type III secretion system (T3SS) is required for virulence in the gram-negative plant pathogen Pseudomonas syringae pv. tomato DC3000. The alternative sigma factor HrpL directly regulates expression of T3SS genes via a promoter sequence, often designated as the “hrp promoter.” Although the HrpL regulon has been extensively investigated in DC3000, it is not known whether additional regulon members remain to be found. To systematically search for HrpL-regulated genes, we used chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-Seq) and bulk mRNA sequencing (RNA-Seq) to identify HrpL-binding sites and likely hrp promoters. The analysis recovered 73 sites of interest, including 20 sites that represent new hrp promoters. The new promoters lie upstream of a diverse set of genes encoding potential regulators, enzymes and hypothetical proteins. PSPTO_5633 is the only new HrpL regulon member that is potentially an effector and is now designated HopBM1. Deletions in several other new regulon members, including PSPTO_5633, PSPTO_0371, PSPTO_2130, PSPTO_2691, PSPTO_2696, PSPTO_3331, and PSPTO_5240, in either DC3000 or ΔhopQ1-1 backgrounds, do not affect the hypersensitive response or in planta growth of the resulting strains. Many new HrpL regulon members appear to be unrelated to the T3SS, and orthologs for some of these can be identified in numerous non-pathogenic bacteria. With the identification of 20 new hrp promoters, the list of HrpL regulon members is approaching saturation and most likely includes all DC3000 effectors.
Highlights
Pseudomonas syringae pv. tomato DC3000 (DC3000), an important model pathogen in molecular plant pathology, causes bacterial speck disease in Arabidopsis [1], tomato [2] and Nicotiana benthamiana (DC3000 mutants lacking virulence determinant HopQ1-1) [3]
Several attempts have been made to define the set of genes regulated by HrpL in DC3000 [21,22,23] and these have succeeded in identifying many effectors and essential components of the T3SS
These efforts were intended to be thorough [24,25], the availability of more advanced methodologies makes it both feasible and worthwhile to conduct a new inventory. The power of this approach was recently demonstrated in a report describing the use of RNA-Seq to link the expression of HrpL to members of the HrpL regulon in six different P. syringae isolates [26]
Summary
Pseudomonas syringae pv. tomato DC3000 (DC3000), an important model pathogen in molecular plant pathology, causes bacterial speck disease in Arabidopsis [1], tomato [2] and Nicotiana benthamiana (DC3000 mutants lacking virulence determinant HopQ1-1) [3]. The T3SS machinery is encoded by the hypersensitive response and pathogenicity (hrp) and hrp conserved (hrc) gene clusters [5]. Effector proteins, encoded by hrp outer protein genes (hop) [6] are translocated into the host cytoplasm via the T3SS to interact with host proteins and/or intervene with host signaling cascades and responses for the benefit of the pathogen [7,8,9]. If one or more effectors or its activities are recognized by the plant immune system (through resistance proteins or other mechanisms), the host hypersensitive response (HR), a localized plant cell death, is triggered and bacterial growth is limited [10]. Effectors are examples of avirulence (avr) genes, a diverse group whose products typically stimulate a strong host defense response
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
