Genome-Wide Identification of Effector Candidates With Conserved Motifs From the Wheat Leaf Rust Fungus Puccinia triticina

Abstract

Rust fungi secrete various specialized effectors into host cells to manipulate the plant defense response. Conserved motifs, including RXLR, LFLAK-HVLVxxP (CRN), Y/F/WxC, CFEM, LysM, EAR, [SG]-P-C-[KR]-P, DPBB_1 (PNPi), and ToxA, have been identified in various oomycete and fungal effectors and are reported to be crucial for effector translocation or function. However, little is known about potential effectors containing any of these conserved motifs in the wheat leaf rust fungus (Puccinia triticina, Pt). In this study, sequencing was performed on RNA samples collected from the germ tubes (GT) of uredospores of an epidemic Pt pathotype PHTT(P) and Pt-infected leaves of a susceptible wheat cultivar "Chinese Spring" at 4, 6, and 8 days postinoculation (dpi). The assembled transcriptome data were compared to the reference genome of "Pt 1-1 BBBD Race 1." A total of 17,976 genes, including 2,284 "novel" transcripts, were annotated. Among all these genes, we identified 3,149 upregulated genes upon Pt infection at all time points compared to GT, whereas 1,613 genes were more highly expressed in GT. A total of 464 secreted proteins were encoded by those upregulated genes, with 79 of them also predicted as possible effectors by EffectorP. Using hmmsearch and Regex, we identified 719 RXLR-like, 19 PNPi-like, 19 CRN-like, 138 Y/F/WxC, and 9 CFEM effector candidates from the deduced protein database including data based on the "Pt 1-1 BBBD Race 1" genome and the transcriptome data collected here. Four of the PNPi-like effector candidates with DPBB_1 conserved domain showed physical interactions with wheat NPR1 protein in yeast two-hybrid assay. Nine Y/F/WxC and seven CFEM effector candidates were transiently expressed in Nicotiana benthamiana. None of these effector candidates showed induction or suppression of cell death triggered by BAX protein, but the expression of one CFEM effector candidate, PTTG_08198, accelerated the progress of cell death and promoted the accumulation of reactive oxygen species (ROS). In conclusion, we profiled genes associated with the infection process of the Pt pathotype PHTT(P). The identified effector candidates with conserved motifs will help guide the investigation of virulent mechanisms of leaf rust fungus.

Transcriptome Analysis Based on Lr19-Virulent Mutants Strain Provides Clues for the Pathogenicity-Related Genes and Effectors of Puccinia Triticina

“…7B). These results were similar with the gene expression patterns predicted based on their FPKM values [28].…”

“…PTTG_25982 was not only a common DEG in M1-vs-WT and M2-vs-WT comparisons, but was also predicted to be an effector candidate. Zhao et al sequenced Pt race PHTT(P) and analyzed the functional factors with conserved motifs [28], in this study, four common genes (PTTG_25271, PTTG_04128, PTTG_08504, and PTTG_08503) encoding PNPi like effectors were found.…”

“…We searched the data of Pt race PHTT(P) [28] and foundthat 6 of 30 effector candidates were differently expressed ( Fig. 7A).…”

Rust effector PNPi interacting with wheat TaPR1a attenuates plant defense response

“…NPR1 may also be a target hub of effectors from various pathogens, including Pseudomonas syringae, Puccinia striiformis f. sp. tritici, and Phytophthora capsici (Wang et al 2016;Chen et al 2017;Li et al 2019;Zhao et al 2020).…”

“…The successful biotrophic lifestyle of rust fungi depends upon the ability of these fungi to deliver specialized effectors into host cells to suppress or evade plant defense. Recent genome sequencing approaches allow the prediction of hypothetical effectors from different wheat rust species (Duplessis et al 2011;Cantu et al 2013a;Zheng et al 2013;Kiran et al 2016;Zhao et al 2020). The few well-characterized rust effector PST02549 from Puccinia striiformis f. sp.…”