Characterization of the Tomato ARF Gene Family Uncovers a Multi-Levels Post-Transcriptional Regulation Including Alternative Splicing

Background: The phytohormone auxin is involved in a wide range of developmental processes and auxin signaling is known to modulate the expression of target genes via two types of transcriptional regulators, namely, Aux/IAA and Auxin Response Factors (ARF). ARFs play a major role in transcriptional activation or repression through direct binding to the promoter of auxin-responsive genes. The present study aims at gaining better insight on distinctive structural and functional features among ARF proteins. Results: Building on the most updated tomato (Solanum lycopersicon) reference genome sequence, a comprehensive set of ARF genes was identified, extending the total number of family members to 22. Upon correction of structural annotation inconsistencies, renaming the tomato ARF family members provided a consensus nomenclature for all ARF genes across plant species. In silico search predicted the presence of putative target site for small interfering RNAs within twelve Sl-ARFs while sequence analysis of the 59-leader sequences revealed the presence of potential small uORF regulatory elements. Functional characterization carried out by transactivation assay partitioned tomato ARFs into repressors and activators of auxin-dependent gene transcription. Expression studies identified tomato ARFs potentially involved in the fruit set process. Genome-wide expression profiling using RNA-seq revealed that at least one third of the gene family members display alternative splicing mode of regulation during the flower to fruit transition. Moreover, the regulation of several tomato ARF genes by both ethylene and auxin, suggests their potential contribution to the convergence mechanism between the signaling pathways of these two hormones. Conclusion: All together, the data bring new insight on the complexity of the expression control of Sl-ARF genes at the transcriptional and post-transcriptional levels supporting the hypothesis that these transcriptional mediators might represent one of the main components that enable auxin to regulate a wide range of physiological processes in a highly specific and coordinated manner

A genomics approach to understanding the role of auxin in apple (Malus x domestica) fruit size control

Background Auxin is an important phytohormone for fleshy fruit development, having been shown to be involved in the initial signal for fertilisation, fruit size through the control of cell division and cell expansion, and ripening related events. There is considerable knowledge of auxin-related genes, mostly from work in model species. With the apple genome now available, it is possible to carry out genomics studies on auxin-related genes to identify genes that may play roles in specific stages of apple fruit development. Results High amounts of auxin in the seed compared with the fruit cortex were observed in 'Royal Gala' apples, with amounts increasing through fruit development. Injection of exogenous auxin into developing apples at the start of cell expansion caused an increase in cell size. An expression analysis screen of auxin-related genes involved in auxin reception, homeostasis, and transcriptional regulation showed complex patterns of expression in each class of gene. Two mapping populations were phenotyped for fruit size over multiple seasons, and multiple quantitative trait loci (QTLs) were observed. One QTL mapped to a region containing an Auxin Response Factor (ARF106). This gene is expressed during cell division and cell expansion stages, consistent with a potential role in the control of fruit size. Conclusions The application of exogenous auxin to apples increased cell expansion, suggesting that endogenous auxin concentrations are at least one of the limiting factors controlling fruit size. The expression analysis of ARF106 linked to a strong QTL for fruit weight suggests that the auxin signal regulating fruit size could partially be modulated through the function of this gene. One class of gene (GH3) removes free auxin by conjugation to amino acids. The lower expression of these GH3 genes during rapid fruit expansion is consistent with the apple maximising auxin concentrations at this point

Data from: QTL analysis of soft scald in two apple populations

The apple (Malus×domestica Borkh.) is one of the world’s most widely grown and valuable fruit crops. With demand for apples year round, storability has emerged as an important consideration for apple breeding programs. Soft scald is a cold storage-related disorder that results in sunken, darkened tissue on the fruit surface. Apple breeders are keen to generate new cultivars that do not suffer from soft scald and can thus be marketed year round. Traditional breeding approaches are protracted and labor intensive, and therefore marker-assisted selection (MAS) is a valuable tool for breeders. To advance MAS for storage disorders in apple, we used genotyping-by-sequencing (GBS) to generate high-density genetic maps in two F1 apple populations, which were then used for quantitative trait locus (QTL) mapping of soft scald. In total, 900 million DNA sequence reads were generated, but after several data filtering steps, only 2% of reads were ultimately used to create two genetic maps that included 1918 and 2818 single-nucleotide polymorphisms. Two QTL associated with soft scald were identified in one of the bi-parental populations originating from parent 11W-12-11, an advanced breeding line. This study demonstrates the utility of next-generation DNA sequencing technologies for QTL mapping in F1 populations, and provides a basis for the advancement of MAS to improve storability of apples

Apple russeting as seen through the RNA-seq lens: strong alterations in the exocarp cell wall

Russeting, a commercially important defect in the exocarp of apple (Malus × domestica), is mainly characterized by the accumulation of suberin on the inner part of the cell wall of the outer epidermal cell layers. However, knowledge on the underlying genetic components triggering this trait remains sketchy. Bulk transcriptomic profiling was performed on the exocarps of three russeted and three waxy apple varieties. This experimental design was chosen to lower the impact of genotype on the obtained results. Validation by qPCR was carried out on representative genes and additional varieties. Gene ontology enrichment revealed a repression of lignin and cuticle biosynthesis genes in russeted exocarps, concomitantly with an enhanced expression of suberin deposition, stress responsive, primary sensing, NAC and MYB-family transcription factors, and specific triterpene biosynthetic genes. Notably, a strong correlation (R2 = 0.976) between the expression of a MYB93-like transcription factor and key suberin biosynthetic genes was found. Our results suggest that russeting is induced by a decreased expression of cuticle biosynthetic genes, leading to a stress response which not only affects suberin deposition, but also the entire structure of the cell wall. The large number of candidate genes identified in this study provides a solid foundation for further functional studies