Orthology Analysis and In Vivo Complementation Studies to Elucidate the Role of DIR1 during Systemic Acquired Resistance in Arabidopsis thaliana and Cucumis sativus

Abstract

AtDIR1 (Defective in Induced Resistance1) is an acidic lipid transfer protein essential for systemic acquired resistance (SAR) in Arabidopsis thaliana. Upon SAR induction, DIR1 moves from locally infected to distant uninfected leaves to activate defense priming; however, a molecular function for DIR1 has not been elucidated. Bioinformatic analysis and in silico homology modeling identified putative AtDIR1 orthologs in crop species, revealing conserved protein motifs within and outside of DIR1’s central hydrophobic cavity. In vitro assays to compare the capacity of recombinant AtDIR1 and targeted AtDIR1-variant proteins to bind the lipophilic probe TNS (6,P-toluidinylnaphthalene-2-sulfonate) provided evidence that conserved leucine 43 and aspartic acid 39 contribute to the size of the DIR1 hydrophobic cavity and possibly hydrophobic ligand binding. An Arabidopsis–cucumber SAR model was developed to investigate the conservation of DIR1 function in cucumber (Cucumis sativus), and we demonstrated that phloem exudates from SAR-induced cucumber rescued the SAR defect in the Arabidopsis dir1-1 mutant. Additionally, an AtDIR1 antibody detected a protein of the same size as AtDIR1 in SAR-induced cucumber phloem exudates, providing evidence that DIR1 function during SAR is conserved in Arabidopsis and cucumber. In vitro TNS displacement assays demonstrated that recombinant AtDIR1 did not bind the SAR signals azelaic acid (AzA), glycerol-3-phosphate or pipecolic acid. However, recombinant CsDIR1 and CsDIR2 interacted weakly with AzA and pipecolic acid. Bioinformatic and functional analyses using the Arabidopsis–cucumber SAR model provide evidence that DIR1 orthologs exist in tobacco, tomato, cucumber, and soybean, and that DIR1-mediated SAR signaling is conserved in Arabidopsis and cucumber.

Sugarcane transcriptome analysis in response to infection caused by Acidovorax avenae subsp. avenae

“…The SAR is characterized by a lasting state of wide spectrum and is normally induced after HR [145], but can also be induced by PTI. Several potential SAR mobile signals have been identified [146]. Numerous studies have shown that DIR is essential for SAR [146–149].…”

“…Several potential SAR mobile signals have been identified [146]. Numerous studies have shown that DIR is essential for SAR [146–149]. Among the DETs it stands out a DIR gene, suggesting induction of SAR in sugarcane infected by Aaa (S8 Table).…”

Signals of Systemic Immunity in Plants: Progress and Open Questions

“…Additional LTPs, like LTP1 and LTP2 and even AZELAIC ACID INDUCED1 (AZI1) and its homologs have a role in SAR signal development [ 54 , 55 ]. Moreover, DIR1 is conserved in other SAR-competent species like cucumber [ 56 ] and tobacco ( Nicotiana tabacum ) [ 57 ]. The lipid/acyl-CoA binding protein, ACBP6 is also required for SAR induction [ 58 ] and present in phloem exudates of SAR induced Arabidopsis plants [ 59 ].…”

The interplay of phloem-mobile signals in plant development and stress response

“…DIR1 is a lipid transfer protein that has been shown to be phloem mobile during SAR and is required for systemic resistance [79,183]. While DIR1 alone does not induce resistance in distant tissues, it may be required for the transport of other SAR signals like DHA [131].…”