Sugar Accumulation in Leaves of Arabidopsis sweet11/sweet12 Double Mutants Enhances Priming of the Salicylic Acid-Mediated Defense Response

Abstract

In compatible interactions, biotrophic microbial phytopathogens rely on the supply of assimilates by the colonized host tissue. It has been found in rice that phloem localized SWEET sucrose transporters can be reprogrammed by bacterial effectors to establish compatibility. We observed that sweet11/sweet12 double mutants, but not single mutants, exhibited increased resistance toward the fungal hemibiotroph Colletotrichum higginsianum (Ch), both in the biotrophic and the necrotrophic colonization phase. We therefore investigated if the phloem localized transporters AtSWEET11 and AtSWEET12 represent additive susceptibility factors in the interaction of Arabidopsis with Ch. AtSWEET12-YFP fusion protein driven by the endogenous promoter strongly accumulated at Ch infection sites and in the vasculature upon challenge with Ch. However, susceptibility of sweet12 single mutants to Ch was comparable to wild type, indicating that the accumulation of AtSWEET12 at Ch infection sites does not play a major role for compatibility. AtSWEET12-YFP reporter protein was not detectable at the plant–pathogen interface, suggesting that AtSWEET12 is not targeted by Ch effectors. AtSWEET11-YFP accumulation in pAtSWEET11:AtSWEET11-YFP plants were similar in Ch infected and mock control leaves. A close inspection of major carbohydrate metabolism in non-infected control plants revealed that soluble sugar and starch content were substantially elevated in sweet11/sweet12 double mutants during the entire diurnal cycle, that diurnal soluble sugar turnover was increased more than twofold in sweet11/sweet12, and that accumulation of free hexoses and sucrose was strongly expedited in double mutant leaves compared to wild type and both single mutants during the course of Ch infection. After 2 days of treatment, free and conjugated SA levels were significantly increased in infected and mock control leaves of sweet11/sweet12 relative to all other genotypes, respectively. Induced genes in mock treated sweet11/sweet12 leaves were highly significantly enriched for several GO terms associated with SA signaling and response compared to mock treated wild-type leaves, indicating sugar-mediated priming of the SA pathway in the double mutant. Infection assays with salicylic acid deficient sweet11/sweet12/sid2 triple mutants demonstrated that reduced susceptibility observed in sweet11/sweet12 was entirely dependent on the SA pathway. We suggest a model how defects in phloem loading of sucrose can influence SA priming and hence, compatibility.

Sugar flux and signaling in plant–microbe interactions

“…STPs, as proton hexose symporters, are known to take up hexoses from apoplasmic space and have been found to be induced during pathogen challenge (Lemonnier et al, 2014), supporting the pathogen starvation hypothesis. On the other hand, sugars themselves can act as signals that induce defense genes (Herbers et al, 1996;Herbers and Sonnewald, 1998;Gebauer et al, 2017), supporting a 'sugar signaling' hypothesis.…”

“…Sonnewald's group put forward a ‘priming’ hypothesis, in which hexose sensing in the secretory pathway mediates the induction of defense genes (Herbers et al ., ). More recently, they showed that defects in phloem loading can trigger SA‐mediated priming of defense during infection of Arabidopsis by Colletotrichum higginsianum , further supporting the ‘sugar signaling’ hypothesis (Gebauer et al ., ).…”

“…This hypothesis depends on the assumption that apoplasmic sugar pools are low, thereby limiting pathogen growth. An alternative is a 'sugar signaling' hypothesis, in which altered levels of sugar at the infection site trigger signaling cascades that result in salicylic acid (SA) pathway activation and defense gene upregulation, ultimately generating physiological changes that repel pathogens (Gebauer et al, 2017).…”

Reactive oxygen species dosage in Arabidopsis chloroplasts can improve resistance towards Colletotrichum higginsianum by the induction of WRKY33

“…Wild‐type Col‐0 and cat2‐2 (SALK_057998; Queval et al ., , ; Sewelam et al ., ), sid2‐3 (SALK_042603; Gebauer et al ., ), and GO5, GO14, GO16, GO20, and GO–MS14 overexpressors (Fahnenstich et al ., ; Sewelam et al ., ) were taken from previous studies. Knockout lines for erf6‐1 (SAIL_1236_H11; Vogel et al ., ) and wrky33‐2 (GABI‐Kat324B11; Mao et al ., ) were obtained from the Nottingham Arabidopsis Stock Centre (UK) and homozygous offspring were identified by PCR; myb34/myb51 (WiscDsLox 424F3/GABI‐KAT228B12; Gigolashvili et al ., ) were donated by Tamara Gigolashivili (University of Cologne, Germany).…”

“…Wild-type Col-0 and cat2-2 (SALK_057998; Queval et al, 2007Queval et al, , 2009Sewelam et al, 2014), sid2-3 (SALK_042603; Gebauer et al, 2017), and GO5, GO14, GO16, GO20, and GO-MS14 overexpressors (Fahnenstich et al, 2008;Sewelam et al, 2014) were taken from previous studies. Knockout lines for erf6-1 (SAIL_1236_H11; Vogel et al, 2014) and wrky33-2 (GABI-Kat324B11; Mao et al, 2011) were obtained from the Nottingham Arabidopsis Stock Centre (UK) and homozygous offspring were identified by PCR; myb34/myb51 (WiscDsLox 424F3/ GABI-KAT228B12; Gigolashvili et al, 2007) Phytologist light : dark,22°C : 19°C) at 50 µmol m À2 s À1 and were then cultivated in a growth cabinet in a 16 h : 8 h, light : dark, cycle at 22°C day : 20°C night temperatures, either under very low light (VLL) conditions with a photon flux density (PFD) of 30 µmol m À2 s À1 , or in low light (LL) conditions at a PFD of 90 µmol m À2 s À1 .…”