A Case of Gene Fragmentation in Plant Mitochondria Fixed by the Selection of a Compensatory Restorer of Fertility-Like PPR Gene.

The high mutational load of mitochondrial genomes combined with their uniparental inheritance and high polyploidy favors the maintenance of deleterious mutations within populations. How cells compose and adapt to the accumulation of disadvantageous mitochondrial alleles remains unclear. Most harmful changes are likely corrected by purifying selection, however, the intimate collaboration between mitochondria- and nuclear-encoded gene products offers theoretical potential for compensatory adaptive changes. In plants, cytoplasmic male sterilities are known examples of nucleo-mitochondrial coadaptation situations in which nuclear-encoded restorer of fertility (Rf) genes evolve to counteract the effect of mitochondria-encoded cytoplasmic male sterility (CMS) genes and restore fertility. Most cloned Rfs belong to a small monophyletic group, comprising 26 pentatricopeptide repeat genes in Arabidopsis, called Rf-like (RFL). In this analysis, we explored the functional diversity of RFL genes in Arabidopsis and found that the RFL8 gene is not related to CMS suppression but essential for plant embryo development. In vitro-rescued rfl8 plantlets are deficient in the production of the mitochondrial heme-lyase complex. A complete ensemble of molecular and genetic analyses allowed us to demonstrate that the RFL8 gene has been selected to permit the translation of the mitochondrial ccmFN2 gene encoding a heme-lyase complex subunit which derives from the split of the ccmFN gene, specifically in Brassicaceae plants. This study represents thus a clear case of nuclear compensation to a lineage-specific mitochondrial genomic rearrangement in plants and demonstrates that RFL genes can be selected in response to other mitochondrial deviancies than CMS suppression.

Redox-mediated kick-start of mitochondrial energy metabolism drives resource-efficient seed germination.

Seeds preserve a far developed plant embryo in a quiescent state. Seed metabolism relies on stored resources and is reactivated to drive germination when the external conditions are favorable. Since the switchover from quiescence to reactivation provides a remarkable case of a cell physiological transition we investigated the earliest events in energy and redox metabolism of Arabidopsis seeds at imbibition. By developing fluorescent protein biosensing in intact seeds, we observed ATP accumulation and oxygen uptake within minutes, indicating rapid activation of mitochondrial respiration, which coincided with a sharp transition from an oxidizing to a more reducing thiol redox environment in the mitochondrial matrix. To identify individual operational protein thiol switches, we captured the fast release of metabolic quiescence in organello and devised quantitative iodoacetyl tandem mass tag (iodoTMT)-based thiol redox proteomics. The redox state across all Cys peptides was shifted toward reduction from 27.1% down to 13.0% oxidized thiol. A large number of Cys peptides (412) were redox switched, representing central pathways of mitochondrial energy metabolism, including the respiratory chain and each enzymatic step of the tricarboxylic acid (TCA) cycle. Active site Cys peptides of glutathione reductase 2, NADPH-thioredoxin reductase a/b, and thioredoxin-o1 showed the strongest responses. Germination of seeds lacking those redox proteins was associated with markedly enhanced respiration and deregulated TCA cycle dynamics suggesting decreased resource efficiency of energy metabolism. Germination in aged seeds was strongly impaired. We identify a global operation of thiol redox switches that is required for optimal usage of energy stores by the mitochondria to drive efficient germination.

Water content, adenylate kinase, and mitochondria drive adenylate balance in dehydrating and imbibing seeds.

Water and life are inexorably linked, but some organisms are capable of losing almost all cellular water to enter a non-metabolic state of anhydrobiosis. This raises intriguing questions about how energy metabolism is managed during such transitions. Here, we have investigated adenylate metabolism during seed imbibition and drying using intact or fragmented pea (Pisum sativum L.) seeds. AMP was confirmed as the major adenylate stored in dry seeds, and normal adenylate balance was rapidly restored upon rehydration of the tissues. Conversely, re-drying of fully imbibed seeds reversed the balance toward AMP accumulation. The overall analysis, supported by in vitro enzyme mimicking experiments, shows that during tissue dehydration, when oxidative phosphorylation is no longer efficient because of decreasing water content, the ATP metabolic demand is met by adenylate kinase, resulting in accumulation of AMP. During seed imbibition, adenylate balance is rapidly restored from the AMP stock by the concerted action of adenylate kinase and mitochondria. The adenylate balance in orthodox seeds, and probably in other anhydrobiotes, appears to be simply driven by water content throughout the interplay between ATP metabolic demand, adenylate kinase, and oxidative phosphorylation, which requires mitochondria to be energetically efficient from the onset of imbibition.

Low oxygen sensing and balancing in plant seeds: a role for nitric oxide.

Storage product accumulation in seeds of major crop species is limited by their low internal oxygen concentration. Adjustment of energy and storage metabolism to oxygen deficiency (hypoxia) in seeds is highly relevant for agriculture and biotechnology. However, the mechanisms of low-oxygen sensing and balancing remain a mystery. Here, it is shown that normal hypoxia in seeds of soybean (Glycine max) and pea (Pisum sativum) triggers a nitrite-dependent increase in endogenous nitric oxide (NO) concentrations. NO, in turn, reduces the oxygen consumption of seeds, generating a localized decrease in both ATP availability and biosynthetic activity. Increasing oxygen availability reduces endogenous NO concentrations, thereby abolishing mitochondrial and metabolic inhibition. This auto-regulatory and reversible oxygen balancing, via NO, avoids seed anoxia and suggests a key role for NO in regulating storage activity. This hypothesis is reinforced by changes in energy status (ATP:ADP ratio), steady-state metabolite concentrations and biosynthetic fluxes under NO treatment. The proposed mechanism of low-oxygen sensing and balancing in plants offers the prospect of a new field of study in crop biotechnology.