Mitochondrial complex II (succinate dehydrogenase [SDH]) is part of the tricarboxylic acid cycle and the respiratory electron transport chain. Its flavoprotein subunit is encoded by two nuclear genes, SDH1-1 and SDH1-2, in Arabidopsis (Arabidopsis thaliana). The SDH1-2 gene is significantly expressed only in roots, albeit at very low level, and its disruption has no effect on growth and development of homozygous mutant plants. In contrast, SDH1-1 transcripts are ubiquitously expressed, with highest expression in flowers. Disruption of the SDH1-1 gene results in alterations in gametophyte development. Indeed, heterozygous SDH1-1/sdh1-1 mutant plants showed normal vegetative growth, yet a reduced seed set. In the progeny of selfed SDH1-1/sdh1-1 plants, distorted segregation ratios were observed, and no homozygous mutant plants were obtained. Reciprocal test crosses with the wild type demonstrated that the mutated sdh1-1 allele is not transmitted through the male gametophyte and is only partially transmitted through the female gametophyte. Consistently, microscopic analysis showed that mutant microspores develop normally until the vacuolated microspore stage, but fail to undergo mitosis I, and then cell structures are degraded and cell content disappears. On the other hand, half the mutant embryo sacs showed arrested development, either at the two-nucleate stage or before polar nuclei fusion. Down-regulation of SDH1-1 by RNA interference results in pollen abortion and a reduced seed set, as in the insertional mutant. Altogether, our results show that SDH1-1, and therefore complex II, are essential for gametophyte development.

RNA editing in higher plant mitochondria modifies mRNA sequences by means of C-to-U conversions at highly specific sites. To determine the cis elements involved in recognition of an editing site in plant mitochondria, deletion and site-directed mutation constructs containing the cognate cox II mitochondrial gene were introduced into purified mitochondria by electroporation. The RNA editing status was analyzed for precursor and spliced transcripts from the test construct. We found that only a restricted number of nucleotides in the vicinity of the target C residue were necessary for recognition by the editing machinery and that the nearest neighbor 3 residues were crucial for the editing process. We provide evidence that two functionally distinguishable sequences can be defined: the 16-nucleotide 5 region, which can be replaced with the same region from another editing site, and a 6-nucleotide 3 region specific to the editing site. The latter region may play a role in positioning the actual editing residue.RNA editing refers to a process whereby the genetic message is changed at single nucleotides in a very specific manner. This process involves a variety of genetic systems and occurs by different mechanisms (reference 6 and references therein). In trypanosome kinetoplasts, RNA editing proceeds by insertion and deletion of uridine nucleotides in mRNAs (2); the insertion of C residues has been described for Physarum polycephalum mitochondria, and the insertion of some G residues occurs in paramyxovirus (25,36). Another type of RNA editing is base conversion, occurring in mammalian nuclei (31) and plant organelles. C-to-U conversions have been described for higher plant mitochondria (9, 13, 16) and to a lesser extent for chloroplasts (18,21).RNA editing is a posttranscriptional event in plant organelles. It is essential in plant mitochondrion gene expression processes such as the maturation step of organellar transcripts (26,27) or the synthesis of functional proteins, since the nucleotide conversions usually alter the coding properties of the mRNA (1). The editing systems in higher plant organelles, mitochondria, and chloroplasts share many similar features, but promiscuous chloroplast sequences are not edited in mitochondria (39); conversely, a mitochondrial sequence carrying an editing site does not sustain editing when transcribed into chloroplasts (35). These results indicate that editing recognition signals are specific to each organelle. The sequences flanking target C residues lack any apparent conserved consensus elements at the primary or secondary structure level. An essential problem is to define the signals that determine the specific recognition of every editing site.A number of in vivo studies of transgenic chloroplasts have demonstrated that mRNA sequences flanking the editing site are involved in RNA editing (4, 5). RNA editing has been determined to proceed by deamination of the C residue in wheat (3) and pea (38). However, the molecular determinants for editing-site recognition have not yet been ...

Three different nuclear genes encode the essential iron-sulfur subunit of mitochondrial complex II (succinate dehydrogenase) in Arabidopsis (Arabidopsis thaliana), raising interesting questions about their origin and function. To find clues about their role, we have undertaken a detailed analysis of their expression. Two genes (SDH2-1 and SDH2-2) that likely arose via a relatively recent duplication event are expressed in all organs from adult plants, whereas transcripts from the third gene (SDH2-3) were not detected. The tissue-and cell-specific expression of SDH2-1 and SDH2-2 was investigated by in situ hybridization. In flowers, both genes are regulated in a similar way. Enhanced expression was observed in floral meristems and sex organ primordia at early stages of development. As flowers develop, SDH2-1 and SDH2-2 transcripts accumulate in anthers, particularly in the tapetum, pollen mother cells, and microspores, in agreement with an essential role of mitochondria during anther development. Interestingly, in contrast to the situation in flowers, only SDH2-2 appears to be expressed at a significant level in root tips. Strong labeling was observed in all cell layers of the root meristematic zone, and a cell-specific pattern of expression was found with increasing distance from the root tip, as cells attain their differentiated state. Analysis of transgenic Arabidopsis plants carrying SDH2-1 and SDH2-2 promoters fused to the b-glucuronidase reporter gene indicate that both promoters have similar activities in flowers, driving enhanced expression in anthers and/or pollen, and that only the SDH2-2 promoter is active in root tips. These b-glucuronidase staining patterns parallel those obtained by in situ hybridization, suggesting transcriptional regulation of these genes. Progressive deletions of the promoters identified regions important for SDH2-1 expression in anthers and/or pollen and for SDH2-2 expression in anthers and/or pollen and root tips. Interestingly, regions driving enhanced expression in anthers are differently located in the two promoters.The mitochondrial electron transport chain of eukaryotes consists of four major multimeric enzyme complexes, one of which is succinate:ubiquinone oxidoreductase (succinate dehydrogenase; EC 1.3.5.1), commonly referred to as complex II. This important membrane-associated complex is a functional part of both the citric acid cycle and the aerobic respiratory chain, catalyzing the oxidation of succinate to fumarate and the reduction of ubiquinone to ubiquinol.Complex II has been well characterized in bacteria, fungi, and mammals and is known to be the simplest of all the complexes of the electron transport chain, with four subunits (Lemire and Oyedotun, 2002;Yankovskaya et al., 2003). It contains two peripheral membrane proteins, a flavoprotein (SDH1) and an iron-sulfur protein (SDH2), and two small integral membrane proteins (SDH3 and SDH4). The succinatebinding site is formed by the SDH1 polypeptide, which is linked covalently to a FAD molecule acting as acceptor o...

Mitochondrial complex II (succinate dehydrogenase) is part of the tricarboxylic acid cycle and the respiratory chain. Three nuclear genes encode its essential iron-sulfur subunit in Arabidopsis (Arabidopsis thaliana). One of them, SUCCINATE DEHYDROGENASE2-3 (SDH2-3), is specifically expressed in the embryo during seed maturation, suggesting that SDH2-3 may have a role as the complex II iron-sulfur subunit during embryo maturation and/or germination. Here, we present data demonstrating that three abscisic acid-responsive elements and one RY-like enhancer element, present in the SDH2-3 promoter, are involved in embryo-specific SDH2-3 transcriptional regulation. Furthermore, we show that ABSCISIC ACID INSENSI-TIVE3 (ABI3), FUSCA3 (FUS3), and LEAFY COTYLEDON2, three key B3 domain transcription factors involved in gene expression during seed maturation, control SDH2-3 expression. Whereas ABI3 and FUS3 interact with the RY element in the SDH2-3 promoter, the abscisic acid-responsive elements are shown to be a target for bZIP53, a member of the basic leucine zipper (bZIP) family of transcription factors. We show that group S1 bZIP53 protein binds the promoter as a heterodimer with group C bZIP10 or bZIP25. To the best of our knowledge, the SDH2-3 promoter is the first embryo-specific promoter characterized for a mitochondrial respiratory complex protein. Characterization of succinate dehydrogenase activity in embryos from two homozygous sdh2-3 mutant lines permits us to conclude that SDH2-3 is the major iron-sulfur subunit of mature embryo complex II. Finally, the absence of SDH2-3 in mutant seeds slows down their germination, pointing to a role of SDH2-3-containing complex II at an early step of germination.