Pathogen induced suppression of ABA enhancesFv/Fm reduction.
ABA biosynthesis and signalling is hijacked by DC3000 to suppress
immunity (de Torres-Zabala et al. , 2007; De Torres Zabalaet al. , 2009). The impact of ABA mutants on virulence is
reflected in Fv/Fm signatures (de
Torres Zabala et al. , 2015). As ABA is made predominately in the
chloroplasts, we investigated whether high light induced susceptibility
was underpinned by ABA signalling. We monitored the impact of an ABA
hypersusceptible signalling mutant (triple mutant) or the ABA
insensitive biosynthetic mutant Arabidopsis aldehyde oxidase 3(aao3 ) on infection under normal and high light, monitoring bothFv/Fm and non-photochemical
quenching (NPQ), the latter measuring the energy released as heat. Theaao3 mutant exhibited less suppression ofFv/Fm during DC3000 infection
compared to Col-0 plants, reflected also in slightly lower levels of NPQ
in comparison to Col-0 (Figure 8A, B, Sup Fig 4a, b). By contrast the
hypersensitive triple PP2C mutant (abi1/abi2/hab1 ) shows a faster
decrease in Fv/Fm and a stronger
increase in NPQ compared to Col-0 (Figure 8A, B, Supp Figure 4a, b). As
previously reported (de Torres-Zabala et al. , 2007; De Torres
Zabala et al. , 2009; Rubio et al. , 2009) under normal
light conditions aao3 plants are more resistant to DC3000 while
the triple PP2C mutant is more susceptible (Figure 8C). However, under
high light (450 µmol m-2s-1) Col-0
and aao3 plants are more susceptible however there was no
enhanced susceptibility evident in the triple PP2C mutant (Figure 8C)
implying either ABA signalling is important for high light enhanced
susceptibility or the abi1/abi2/hab1 plants cannot support
further bacterial multiplication. In addition, Col-0 plants grown under
high light show accumulation of ABA after 5 days of high light growth
and 9 days of high light with subsequent DC3000 infection compared to no
increase in ABA under normal light or bacterial infection (Supp Figure
3c). In contrast the aao3 plants do not show an increase in ABA under
normal or high light (Supp Figure 3c).
To next assess the interaction of ABA and light on chloroplast function
during pathogen infection 10 µM ABA was co-infiltrated with DC3000 into
Col-0 leaves. Under normal light, 10 µM ABA co-infiltration enhances the
decrease in Fv/Fm levels as
previously reported (de Torres Zabala et al. , 2015)(Figure 8D,
E). Under high light conditions infiltration with 10 µM and 100 µM ABA
treatments show a faster decrease ofFv/Fm levels from 3.5 hours
onwards (Figure 8F, G). To ensure that 10 and 100 µM ABA treatment was
not toxic to P. syringae , DC3000 was plated on Kings B agar
containing 0, 10, 50 and 100 µM ABA. Bacterial growth showed that there
is a small reduction in growth of bacteria in the presence of ABA,
notably being significant (p<0.0005)with increased ABA
concentration, (Supp Figure 4c).