MISF2 Encodes an Essential Mitochondrial Splicing Co-factor Required for nad2 mRNA Processing and Embryo Development in Arabidopsis thaliana

Mitochondria play key roles in cellular energy metabolism in eukaryotes. Mitochondria of most organisms contain their own genome and specific transcription and translation machineries. The expression of angiosperm mtDNA involves extensive RNA-processing steps, such as RNA trimming, editing, and the splicing of numerous group II-type introns. Pentatricopeptide repeat (PPR) proteins are key players of plant organelle gene expression and RNA metabolism. In the present analysis, we reveal the function of the MITOCHONDRIAL SPLICING FACTOR 2 gene (MISF2, AT3G22670) and show that it encodes a mitochondria-localized PPR protein that is crucial for early embryo-development in Arabidopsis. Molecular characterization of embryo-rescued misf2 plantlets indicates that the splicing of nad2 intron 1 and thus respiratory complex I biogenesis are strongly compromised. Moreover, the molecular function seems conserved between MISF2 protein in Arabidopsis and its orthologous gene (EMP10) in maize, suggesting that the ancestor of MISF2/EMP10 was recruited to function in nad2 processing before the monocot-dicot divergence, ~200 million years ago. These data provide new insights into the function of nuclear-encoded factors in mitochondrial gene expression and respiratory chain biogenesis during plant embryo development.

Flowering plant embryos: How did we end up here?

The seeds of flowering plants are sexually produced propagules that ensure dispersal and resilience of the next generation. Seeds harbor embryos, three dimensional structures that are often miniatures of the adult plant in terms of general structure and primordial organs. In addition, embryos contain the meristems that give rise to post-embryonically generated structures. However common, flowering plant embryos are an evolutionary derived state. Flowering plants are part of a much larger group of embryo-bearing plants, aptly termed Embryophyta. A key question is what evolutionary trajectory led to the emergence of flowering plant embryos. In this opinion, we deconstruct the flowering plant embryo and describe the current state of knowledge of embryos in other plant lineages. While we are far yet from understanding the ancestral state of plant embryogenesis, we argue what current knowledge may suggest and how the knowledge gaps may be closed.

LARVAL STRATEGY OF TWO SPECIES OF SEED-FEEDING CHALCIDOIDEA PARALLELS THAT OF PARASITOID KOINOBIONTS

The association between ontogenetic processes in plants and phytophagous insects is not traditionally considered in studies of insect-plant interactions. Angiosperm seeds impose important constraints on seed predators; the structural complexity of seeds and the progressive accumulation of resources throughout their development limit the time windows when resources can be retrieved by the predator. Some holometabolous insects deposit their eggs inside immature seeds, with the immature stages of both insect and plant cohabiting in a space with limited but potential resources. We studied the larval development of Megastigmus transvaalensis (Hussey, 1956) (Chalcidoidea: Megastigmidae) and Bephratelloides pomorum (F., 1804) (Chalcidoidea: Eurytomidae) and the seed development of their respective hosts, Schinus terebinthifolia (Raddi, 1820) (Anacardiaceae) and Annona crassiflora (Mart., 1841) (Annonaceae). Our results show that both M. transvaalensis and B. pomorum oviposit in immature fruits, whose protective tissues surrounding the seeds are softer. The first larval instar interferes little with the development of the seed, allowing both seed and plant embryo to continue growing. When the infested seed reaches the size of a mature, non-infested seed, the larva grows rapidly and consumes most of it. M. transvaalensis induces minor modifications in the endosperm cells, while B. pomorum does not induce any visual modifications. The strategy of allowing seed/plant embryo to continue growing shows similarities to the endoparasitic koinobiont strategy followed by some chalcid parasitoids, which keep their host alive while feeding upon it. Future studies should be expanded to other chalcid seed predators in order to understand the evolution of convergent patterns among seed-feeding insects and its evolution in relation to parasitoid strategies inside the group

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 ofArabidopsisseeds 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.

Redox-mediated Kick-Start of Mitochondrial Energy Metabolism drives Resource-efficient Seed Germination

ABSTRACTSeeds preserve a far developed plant embryo in a quiescent state. Seed metabolism relies on stored resources and is re-activated to drive germination when the external conditions are favorable. Since the switchover from quiescence to re-activation provides a remarkable case of a cell physiological transition we investigated the earliest events in energy and redox metabolism ofArabidopsisseeds 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 quiescencein organelloand devised quantitative iodoacetyl tandem mass tag-based (iodoTMT) thiol redox proteomics. The redox state across all Cys-peptides was shifted towards reduction from 27.1 % to 13.0 %. 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 cycle (TCA). 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.

A shared cis-regulatory module activates transcription in the suspensor of plant embryos

The mechanisms controlling the transcription of gene sets in specific regions of a plant embryo shortly after fertilization remain unknown. Previously, we showed that G564 mRNA, encoding a protein of unknown function, accumulates to high levels in the giant suspensor of both Scarlet Runner Bean (SRB) and Common Bean embryos, and a cis-regulatory module containing three unique DNA sequences, designated as the 10-bp, Region 2, and Fifth motifs, is required for G564 suspensor-specific transcription [Henry KF, et al. (2015) Plant Mol Biol 88:207–217; Kawashima T, et al. (2009) Proc Natl Acad Sci USA 106:3627–3632]. We tested the hypothesis that these motifs are also required for transcription of the SRB GA 20-oxidase gene, which encodes a gibberellic acid hormone biosynthesis enzyme and is coexpressed with G564 at a high level in giant bean suspensors. We used deletion and gain-of-function experiments in transgenic tobacco embryos to show that two GA 20-oxidase DNA regions are required for suspensor-specific transcription, one in the 5′ UTR (+119 to +205) and another in the 5′ upstream region (−341 to −316). Mutagenesis of sequences in these two regions determined that the cis-regulatory motifs required for G564 suspensor transcription are also required for GA 20-oxidase transcription within the suspensor, although the motif arrangement differs. Our results demonstrate the flexibility of motif positioning within a cis-regulatory module that activates gene transcription within giant bean suspensors and suggest that G564 and GA 20-oxidase comprise part of a suspensor gene regulatory network.