Growth Developmental Defects of Mitochondrial Iron Transporter 1 and 2 Mutants in Arabidopsis in Iron Sufficient Conditions.

Iron is the most abundant micronutrient in plant mitochondria, and it has a crucial role in biochemical reactions involving electron transfer. It has been described in that () is an essential gene and that knockdown mutant rice plants have a decreased amount of iron in their mitochondria, strongly suggesting that OsMIT is involved in mitochondrial iron uptake. In , two genes encode MIT homologues. In this study, we analyzed different and mutant alleles, and no phenotypic defects were observed in individual mutant plants grown in normal conditions, confirming that neither nor are individually essential. When we generated crosses between the and alleles, we were able to isolate homozygous double mutant plants. Interestingly, homozygous double mutant plants were obtained only when mutant alleles of with the T-DNA insertion in the intron region were used for crossings, and in these cases, a correctly spliced mRNA was generated, although at a low level. double homozygous mutant plants, knockout for and knockdown for , were grown and characterized in iron-sufficient conditions. Pleiotropic developmental defects were observed, including abnormal seeds, an increased number of cotyledons, a slow growth rate, pinoid stems, defects in flower structures, and reduced seed set. A RNA-Seq study was performed, and we could identify more than 760 genes differentially expressed in . Our results show that double homozygous mutant plants misregulate genes involved in iron transport, coumarin metabolism, hormone metabolism, root development, and stress-related response. The phenotypes observed, such as pinoid stems and fused cotyledons, in double homozygous mutant plants may suggest defects in auxin homeostasis. Unexpectedly, we observed a possible phenomenon of T-DNA suppression in the next generation of double homozygous mutant plants, correlating with increased splicing of the A intron containing the T-DNA and the suppression of the phenotypes observed in the first generation of the double mutant plants. In these plants with a suppressed phenotype, no differences were observed in the oxygen consumption rate of isolated mitochondria; however, the molecular analysis of gene expression markers, , , and , for mitochondrial and oxidative stress showed that these plants express a degree of mitochondrial perturbation. Finally, we could establish by a targeted proteomic analysis that a protein level of 30% of MIT2, in the absence of MIT1, is enough for normal plant growth under iron-sufficient conditions.