Effects of autophagy and CV deficiency on metabolic adjustment
under extended darkness
To gain more insight into the differential impacts of CV and autophagy
deficiency under extended darkness, we further analyzed the metabolic
profile of amircv1xatg5 along with its parental lines,amircv-1 and atg5 (Fig. 6 and 7). In agreement with the
phenotype observed, amircv-1 presented similar metabolic changes
to WT plants, while atg5 and amircv1xatg5 exhibited more
comparable responses. While the majority of amino acids were highly
accumulated in WT and amircv-1 plants, only minor accumulations
of arginine, glutamate, glycine, isoleucine, lysine, serine, threonine
and valine were observed in atg5 and amircvxatg5 after 10d
of extended darkness (Fig. 6). We also found that the TCA cycle
intermediates α-ketoglutarate, citrate, fumarate, malate, and succinate
were highly accumulated in both atg5 and amircvxatg5mutants after 10d of darkness (Fig. 7). These results are consistent
with previous findings demonstrating the impairment of amino acid
provision coupled with an altered respiratory response in atgmutants under extended darkness (Barros et al., 2017; Hirota et al.,
2018).
Despite the similarities with the atg5 mutant,amircv1xatg5 also showed specific metabolic signatures. The
general accumulation of organic acids was lower in theamircv1xatg5 mutant compared to the atg5 mutant after 10d
of darkness (Fig. 7). Additionally, the amircv1xatg5 mutant
displayed lower accumulation of arginine, aspartate, glutamate,
isoleucine, serine and valine levels in the later stages of darkness
(Fig. 6). Glutamate, glutamine, asparagine, aspartate are important
components of nitrogen assimilation, recycling, transport and storage in
plants (Gaufichon et al., 2010). The coordination between CV and
nitrogen assimilation pathways was previously proposed during water
stress and elevated CO2 conditions wherein rice CV
mutants presented altered levels of amino acids (Sade et al., 2018;
Umnajkitikorn et al., 2020). On the other hand, the disruption of
autophagy impacts the supply of amino acids and respiratory rates under
carbon depletion (Izumi et al., 2013; Avin-Wittenberg et al., 2015;
Barros et al., 2017; Hirota et al., 2018). Collectively, our results
indicate that a deficiency solely of CV has only minor effects on plant
responses to extended darkness, and suggests that the metabolic
reprogramming of atg5 mutants under extended darkness is, at
least partially, dependent on the operation of a functional CV pathway.