Oh, G. G. K. | Search bioRxiv

bioRxiv2022

Knockdown of mitochondrial atp1 mRNA by a custom-designed pentatricopeptide repeat protein alters F1Fo ATP synthase

We show that a custom-designed RNA-binding protein binds and specifically induces cleavage of atp1 RNA in mitochondria, significantly decreasing the abundance of the Atp1 protein and the assembled F1Fo ATP synthase in Arabidopsis thaliana. The transformed plants are characterized by delayed vegetative growth and reduced fertility. Five-fold depletion of Atp1 level was accompanied by a decrease in abundance of other ATP synthase subunits, lowered ATP synthesis rate of isolated mitochondria, but no change to mitochondrial electron transport chain complexes, adenylates or energy charge in planta. Transcripts for amino acid transport and a variety of stress response processes were differentially expressed in lines containing the PPR protein, indicating changes to achieve cellular homeostasis when ATP synthase was highly depleted. Leaves of ATP-synthase-depleted lines showed higher respiratory rates and elevated levels of most amino acids at night, most notably serine family amino acids. The results show the value of using custom-designed PPR proteins to influence expression of specific mitochondrial transcripts to carry out reverse genetics studies on mitochondrial gene functions and the consequences of ATP synthase depletion on cellular functions in Arabidopsis. One sentence SummaryKnockdown of mitochondrial atp1 mRNA by a custom-designed pentatricopeptide repeat protein alters F1Fo ATP synthase, plant growth and amino acid metabolism and ATP synthesis in Arabidopsis thaliana

plant biology↗

bioRxiv2021

Alternative oxidase1a and 1d limit proline-dependent oxidative stress and aid salinity recovery in Arabidopsis

A link between Pro catabolism and mitochondrial reactive oxygen species production has been established across eukaryotes and in plants increases in leaf respiration rates have been reported following Pro exposure. Here we investigated how alternative oxidases (AOXs) of the mitochondrial electron transport chain accommodate the large, atypical flux resulting from Pro catabolism and limit oxidative stress during Pro breakdown in mature Arabidopsis leaves. Following Pro treatment, AOX1a and AOX1d accumulate at transcript and protein levels, with AOX1d approaching the level of the typically dominant AOX1a isoform. We therefore sought to determine the function of both AOX isoforms under Pro respiring conditions. Oxygen consumption rate measurements in aox1a and aox1d leaves suggested these AOXs can functionally compensate for each other to establish enhanced AOX catalytic capacity in response to Pro. Generation of aox1a.aox1d lines showed complete loss of AOX proteins and activity upon Pro treatment, yet full respiratory induction in response to Pro remained possible via the cytochrome pathway. However, aox1a.aox1d leaves suffered increased levels of oxidative stress and damage during Pro metabolism compared to WT or the single mutants. During recovery from salt stress, when high rates of Pro catabolism occur naturally, photosynthetic rates in aox1a.aox1d recovered slower than WT or the single aox lines, showing that both AOX1a and AOX1d are beneficial for cellular metabolism during Pro drawdown following osmotic stress. This work provides physiological evidence of a beneficial role for AOX1a but also the less studied AOX1d isoform in allowing safe catabolism of alternative respiratory substrates like Pro. One sentence summaryThe alternative oxidase of plant mitochondria contributes to Pro catabolism by preventing oxidative stress in the electron transport chain and this aids recovery of leaf metabolic rates following salinity stress.

plant biology↗