Abstract
Cell wall reinforcement with callose is a frequent plant response to infection. Poly(ADP-ribosyl)ation is a protein post-translational modification mediated by poly(ADP-ribose) polymerases (PARPs). Poly(ADP-ribosyl)ation has well-known roles in DNA damage repair and has more recently been shown to contribute to plant immune responses. 3-aminobenzamide (3AB) is an established PARP inhibitor and it blocks the callose deposition elicited by flg22 or elf18, two microbe-associated molecular patterns (MAMPs). However, we report that an Arabidopsis parp1parp2parp3 triple mutant does not exhibit loss of flg22-induced callose deposition. Additionally, the more specific PARP inhibitors PJ-34 and INH2BP inhibit PARP activity in Arabidopsis but do not block MAMP-induced callose deposition. These data demonstrate off-target activity of 3AB and indicate that 3AB inhibits callose deposition through a mechanism other than poly(ADP-ribosyl)ation. POWDERY MILDEW RESISTANT 4 (PMR4) is the callose synthase responsible for the majority of MAMP- and wound-induced callose deposition in Arabidopsis. 3AB does not block wound-induced callose deposition, and 3AB does not reduce the PMR4 mRNA abundance increase in response to flg22. Levels of PMR4-HA protein increase in response to flg22, and increase even more in flg22 + 3AB despite no callose being produced. The callose synthase inhibitor 2-deoxy-D-glucose does not cause similar impacts on PMR4-HA protein levels. Beyond MAMPs, we find that 3AB also reduces callose deposition induced by powdery mildew (Golovinomyces cichoracearum) and impairs the penetration resistance of a PMR4 overexpression line. 3AB thus reveals pathogenesis-associated pathways that activate callose synthase enzymatic activity distinct from those that elevate PMR4 mRNA and protein abundance.
Highlights
Plants face numerous potential pathogen invaders, yet they are able to effectively prevent the large majority of these encounters from progressing to disease
3AB Blocks flg22-Induced Callose but Not Wound-Induced Callose In Arabidopsis, flg22 and wound-induced callose are the product of the same POWDERY MILDEW RESISTANT 4 (PMR4) callose synthase enzyme, as both are absent in a pmr4 mutant (Jacobs et al, 2003; Nishimura et al, 2003; Luna et al, 2011)
Even within a single seedling exposed to flg22 and with one cotyledon wounded, we found that 3AB blocks the flg22induced callose but not the wound-induced callose (Figure 1)
Summary
Plants face numerous potential pathogen invaders, yet they are able to effectively prevent the large majority of these encounters from progressing to disease. Plants have evolved pattern recognition receptors (PRRs) that induce defense responses after recognizing certain characteristic compounds of microbes, known as microbe- or pathogen-associated molecular patterns (MAMPs, or PAMPs) (Jones and Dangl, 2006; Boller and Felix, 2009). PTI involves a variety of plant defense responses, including deposition of the β-(1,3)-glucan callose at plant cell walls (Dodds and Rathjen, 2010; Macho and Zipfel, 2014; Li et al, 2016). Callose serves a variety of specialized functions in the cell walls of higher plants. PMR4 (POWDERY MILDEW RESISTANT 4; known as GLUCAN SYNTHASE-LIKE 5 [GSL5] and CALS12) accounts for most wound- and pathogen-induced callose, as both responses are absent in pmr mutants (Jacobs et al, 2003; Nishimura et al, 2003; Clay et al, 2009; Luna et al, 2011)
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