Mitogenomic Research of Silverleaf Sunflower (Helianthus argophyllus) and Its Interspecific Hybrids.

Interspecific hybridization is widespread for sunflowers, both in wild populations and commercial breeding. One of the most common species that can efficiently cross with Helianthus annuus is the Silverleaf sunflower-Helianthus argophyllus. The current study carried out structural and functional organization analyses of mitochondrial DNA in H. argophyllus and the interspecific hybrid, H. annuus (VIR114A line) × H. argophyllus. The complete mitogenome of H. argophyllus counts 300,843 bp, has a similar organization to the mitogenome of cultivated sunflower, and holds SNPs typical for wild sunflowers. RNA editing analysis predicted 484 sites in H. argophyllus mitochondrial CDS. The mitochondrial genome of the H. annuus × H. argophyllus hybrid is identical to the maternal line (VIR114A). We expected that significant rearrangements in the mitochondrial DNA of the hybrid would occur, due to the frequent recombination. However, the hybrid mitogenome lacks rearrangements, presumably due to the preservation of nuclear-cytoplasmic interaction paths.

The Insights into Mitochondrial Genomes of Sunflowers.

The significant difference in the mtDNA size and structure with simultaneous slow evolving genes makes the mitochondrial genome paradoxical among all three DNA carriers in the plant cell. Such features make mitochondrial genome investigations of particular interest. The genus Helianthus is a diverse taxonomic group, including at least two economically valuable species-common sunflower (H. annuus) and Jerusalem artichoke (H. tuberosus). The successful investigation of the sunflower nuclear genome provided insights into some genomics aspects and significantly intensified sunflower genetic studies. However, the investigations of organelles' genetic information in Helianthus, especially devoted to mitochondrial genomics, are presented by limited studies. Using NGS sequencing, we assembled the complete mitochondrial genomes for H. occidentalis (281,175 bp) and H. tuberosus (281,287 bp) in the current investigation. Besides the master circle chromosome, in the case of H. tuberosus, the 1361 bp circular plasmid was identified. The mitochondrial gene content was found to be identical for both sunflower species, counting 32 protein-coding genes, 3 rRNA, 23 tRNA genes, and 18 ORFs. The comparative analysis between perennial sunflowers revealed common and polymorphic SSR and SNPs. Comparison of perennial sunflowers with H. annuus allowed us to establish similar rearrangements in mitogenomes, which have possibly been inherited from a common ancestor after the divergence of annual and perennial sunflower species. It is notable that H. occidentalis and H. tuberosus mitogenomes are much more similar to H. strumosus than H. grosseserratus.

Genotyping and lipid profiling of 601 cultivated sunflower lines reveals novel genetic determinants of oil fatty acid content.

BACKGROUND: Sunflower is an important oilseed crop domesticated in North America approximately 4000 years ago. During the last century, oil content in sunflower was under strong selection. Further improvement of oil properties achieved by modulating its fatty acid composition is one of the main directions in modern oilseed crop breeding. RESULTS: We searched for the genetic basis of fatty acid content variation by genotyping 601 inbred sunflower lines and assessing their lipid and fatty acid composition. Our genome-wide association analysis based on the genotypes for 15,483 SNPs and the concentrations of 23 fatty acids, including minor fatty acids, revealed significant genetic associations for eleven of them. Identified genomic regions included the loci involved in rare fatty acids variation on chromosomes 3 and 14, explaining up to 34.5% of the total variation of docosanoic acid (22:0) in sunflower oil. CONCLUSIONS: This is the first large scale implementation of high-throughput lipidomic profiling to sunflower germplasm characterization. This study contributes to the genetic characterization of Russian sunflower collections, which made a substantial contribution to the development of sunflower as the oilseed crop worldwide, and provides new insights into the genetic control of oil composition that can be implemented in future studies.

Organization Features of the Mitochondrial Genome of Sunflower (Helianthus annuus L.) with ANN2-Type Male-Sterile Cytoplasm.

This study provides insights into the flexibility of the mitochondrial genome in sunflower (Helianthus annuus L.) as well as into the causes of ANN2-type cytoplasmic male sterility (CMS). De novo assembly of the mitochondrial genome of male-sterile HA89(ANN2) sunflower line was performed using high-throughput sequencing technologies. Analysis of CMS ANN2 mitochondrial DNA sequence revealed the following reorganization events: twelve rearrangements, seven insertions, and nine deletions. Comparisons of coding sequences from the male-sterile line with the male-fertile line identified a deletion of orf777 and seven new transcriptionally active open reading frames (ORFs): orf324, orf327, orf345, orf558, orf891, orf933, orf1197. Three of these ORFs represent chimeric genes involving atp6 (orf1197), cox2 (orf558), and nad6 (orf891). In addition, orf558, orf891, orf1197, as well as orf933, encode proteins containing membrane domain(s), making them the most likely candidate genes for CMS development in ANN2. Although the investigated CMS phenotype may be caused by simultaneous action of several candidate genes, we assume that orf1197 plays a major role in developing male sterility in ANN2. Comparative analysis of mitogenome organization in sunflower lines representing different CMS sources also allowed identification of reorganization hot spots in the mitochondrial genome of sunflower.

Characterization of the mitochondrial genome of the MAX1 type of cytoplasmic male-sterile sunflower.

BACKGROUND: More than 70 cytoplasmic male sterility (CMS) types have been identified in Helianthus, but only for less than half of them, research of mitochondrial organization has been conducted. Moreover, complete mitochondrion sequences have only been published for two CMS sources - PET1 and PET2. It has been demonstrated that other sunflower CMS sources like MAX1, significantly differ from the PET1 and PET2 types. However, possible molecular causes for the CMS induction by MAX1 have not yet been proposed. In the present study, we have investigated structural changes in the mitochondrial genome of HA89 (MAX1) CMS sunflower line in comparison to the fertile mitochondrial genome. RESULTS: Eight significant major reorganization events have been determined in HA89 (MAX1) mtDNA: one 110 kb inverted region, four deletions of 439 bp, 978 bp, 3183 bp and 14,296 bp, respectively, and three insertions of 1999 bp, 5272 bp and 6583 bp. The rearrangements have led to functional changes in the mitochondrial genome of HA89 (MAX1) resulting in the complete elimination of orf777 and the appearance of new ORFs - orf306, orf480, orf645 and orf1287. Aligning the mtDNA of the CMS sources PET1 and PET2 with MAX1 we found some common reorganization features in their mitochondrial genome sequences. CONCLUSION: The new open reading frame orf1287, representing a chimeric atp6 gene, may play a key role in MAX1 CMS phenotype formation in sunflower, while the contribution of other mitochondrial reorganizations seems to appear negligible for the CMS development.

Mitochondrial genomes organization in alloplasmic lines of sunflower (Helianthus annuus L.) with various types of cytoplasmic male sterility.

BACKGROUND: Cytoplasmic male sterility (CMS) is a common phenotype in higher plants, that is often associated with rearrangements in mitochondrial DNA (mtDNA), and is widely used to produce hybrid seeds in a variety of valuable crop species. Investigation of the CMS phenomenon promotes understanding of fundamental issues of nuclear-cytoplasmic interactions in the ontogeny of higher plants. In the present study, we analyzed the structural changes in mitochondrial genomes of three alloplasmic lines of sunflower (Helianthus annuus L.). The investigation was focused on CMS line PET2, as there are very few reports about its mtDNA organization. METHODS: The NGS sequencing, de novo assembly, and annotation of sunflower mitochondrial genomes were performed. The comparative analysis of mtDNA of HA89 fertile line and two HA89 CMS lines (PET1, PET2) occurred. RESULTS: The mtDNA of the HA89 fertile line was almost identical to the HA412 line (NC_023337). The comparative analysis of HA89 fertile and CMS (PET1) analog mitochondrial genomes revealed 11,852 bp inversion, 4,732 bp insertion, 451 bp deletion and 18 variant sites. In the mtDNA of HA89 (PET2) CMS line we determined 27.5 kb and 106.5 kb translocations, 711 bp and 3,780 bp deletions, as well as, 5,050 bp and 15,885 bp insertions. There are also 83 polymorphic sites in the PET2 mitochondrial genome, as compared with the fertile line. DISCUSSION: The observed mitochondrial reorganizations in PET1 resulted in only one new open reading frame formation (orfH522), and PET2 mtDNA rearrangements led to the elimination of orf777, duplication of atp6 gene and appearance of four new ORFs with transcription activity specific for the HA89 (PET2) CMS line-orf645, orf2565, orf228 and orf285. Orf228 and orf285 are the atp9 chimeric ORFs, containing transmembrane domains and possibly may impact on mitochondrial membrane potential. So orf228 and orf285 may be the cause for the appearance of the PET2 CMS phenotype, while the contribution of other mtDNA reorganizations in CMS formation is negligible.