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Pepino (Solanum muricatum) is an herbaceous crop phylogenetically related to tomato and potato. Pepino fruit vary in color, size and shape, and are eaten fresh. In this study, we use pepino as a fruit model to understand the transcriptional regulatory mechanisms controlling fruit quality. To identify the key genes involved in anthocyanin biosynthesis in pepino, two genotypes were studied that contrasted in foliar and fruit pigmentation. Anthocyanin profiles were analyzed, as well as the expression of genes that encode enzymes for anthocyanin biosynthesis and transcriptional regulators using both RNA-seq and quantitative PCR. The differential expression of the transcription factor genes R2R3 MYB SmuMYB113 and R3MYB SmuATV suggested their association with purple skin and foliage phenotype. Functional analysis of these genes in both tobacco and pepino showed that SmuMYB113 activates anthocyanins, while SmuATV suppresses anthocyanin accumulation. However, despite elevated expression in all tissues, SmuMYB113 does not significantly elevate flesh pigmentation, suggesting a strong repressive background in fruit flesh tissue. These results will aid understanding of the differential regulation controlling fruit quality aspects between skin and flesh in other fruiting species. [ABSTRACT FROM AUTHOR]

Summary: Phosphatidylethanolamine‐binding protein (PEBP) genes regulate flowering and architecture in many plant species. Here, we study kiwifruit (Actinidia chinensis, Ac) PEBP genes with homology to BROTHER OF FT AND TFL1 (BFT). CRISPR‐Cas9 was used to target AcBFT genes in wild‐type and fast‐flowering kiwifruit backgrounds. The editing construct was designed to preferentially target AcBFT2, whose expression is elevated in dormant buds. Acbft lines displayed an evergrowing phenotype and increased branching, while control plants established winter dormancy. The evergrowing phenotype, encompassing delayed budset and advanced budbreak after defoliation, was identified in multiple independent lines with edits in both alleles of AcBFT2. RNA‐seq analyses conducted using buds from gene‐edited and control lines indicated that Acbft evergrowing plants had a transcriptome similar to that of actively growing wild‐type plants, rather than dormant controls. Mutations in both alleles of AcBFT2 did not promote flowering in wild‐type or affect flowering time, morphology and fertility in fast‐flowering transgenic kiwifruit. In summary, editing of AcBFT2 has the potential to reduce plant dormancy with no adverse effect on flowering, giving rise to cultivars better suited for a changing climate. [ABSTRACT FROM AUTHOR]

Summary: Temperate perennials require exposure to chilling temperatures to resume growth in the following spring. Growth and dormancy cycles are controlled by complex genetic regulatory networks and are governed by epigenetic mechanisms, but the specific genes and mechanisms remain poorly understood.To understand how seasonal changes and chilling regulate dormancy and growth in the woody perennial vine kiwifruit (Ac, Actinidia chinensis), a transcriptome study of kiwifruit buds in the field and controlled conditions was performed. A MADS‐box gene with homology to Arabidopsis FLOWERING LOCUS C (FLC) was identified and characterized.Elevated expression of AcFLC‐like (AcFLCL) was detected during bud dormancy and chilling. A long noncoding (lnc) antisense transcript with an expression pattern opposite to AcFLCL and shorter sense noncoding RNAs were identified. Chilling induced an increase in trimethylation of lysine‐4 of histone H3 (H3K4me3) in the 5′ end of the gene, indicating multiple layers of epigenetic regulation in response to cold. Overexpression of AcFLCL in kiwifruit gave rise to plants with earlier budbreak, whilst gene editing using CRISPR‐Cas9 resulted in transgenic lines with substantially delayed budbreak, suggesting a role in activation of growth.These results have implications for the future management and breeding of perennials for resilience to changing climate. [ABSTRACT FROM AUTHOR]

Abstract Background Flower development in kiwifruit (Actinidia spp.) is initiated in the first growing season, when undifferentiated primordia are established in latent shoot buds. These primordia can differentiate into flowers in the second growing season, after the winter dormancy period and upon accumulation of adequate winter chilling. Kiwifruit is an important horticultural crop, yet little is known about the molecular regulation of flower development. Results To study kiwifruit flower development, nine MADS-box genes were identified and functionally characterized. Protein sequence alignment, phenotypes obtained upon overexpression in Arabidopsis and expression patterns suggest that the identified genes are required for floral meristem and floral organ specification. Their role during budbreak and flower development was studied. A spontaneous kiwifruit mutant was utilized to correlate the extended expression domains of these flowering genes with abnormal floral development. Conclusions This study provides a description of flower development in kiwifruit at the molecular level. It has identified markers for flower development, and candidates for manipulation of kiwifruit growth, phase change and time of flowering. The expression in normal and aberrant flowers provided a model for kiwifruit flower development.

Abstract Background Plant microRNAs (miRNAs) are a class of small, non-coding RNAs that play an important role in development and environmental responses. Hundreds of plant miRNAs have been identified to date, mainly from the model species for which there are available genome sequences. The current challenge is to characterise miRNAs from plant species with agricultural and horticultural importance, to aid our understanding of important regulatory mechanisms in crop species and enable improvement of crops and rootstocks. Results Based on the knowledge that many miRNAs occur in large gene families and are highly conserved among distantly related species, we analysed expression of twenty-one miRNA sequences in different tissues of apple (Malus x domestica 'Royal Gala'). We identified eighteen sequences that are expressed in at least one of the tissues tested. Some, but not all, miRNAs expressed in apple tissues including the phloem tissue were also detected in the phloem sap sample derived from the stylets of woolly apple aphids. Most of the miRNAs detected in apple phloem sap were also abundant in the phloem sap of herbaceous species. Potential targets for apple miRNAs were identified that encode putative proteins shown to be targets of corresponding miRNAs in a number of plant species. Expression patterns of potential targets were analysed and correlated with expression of corresponding miRNAs. Conclusions This study validated tissue-specific expression of apple miRNAs that target genes responsible for plant growth, development, and stress response. A subset of characterised miRNAs was also present in the apple phloem translocation stream. A comparative analysis of phloem miRNAs in herbaceous species and woody perennials will aid our understanding of non-cell autonomous roles of miRNAs in plants.

Abstract MicroRNAs (miRNAs) are a class of small non-coding RNAs with a critical role in development and environmental responses. Efficient and reliable detection of miRNAs is an essential step towards understanding their roles in specific cells and tissues. However, gel-based assays currently used to detect miRNAs are very limited in terms of throughput, sensitivity and specificity. Here we provide protocols for detection and quantification of miRNAs by RT-PCR. We describe an end-point and real-time looped RT-PCR procedure and demonstrate detection of miRNAs from as little as 20 pg of plant tissue total RNA and from total RNA isolated from as little as 0.1 μl of phloem sap. In addition, we have developed an alternative real-time PCR assay that can further improve specificity when detecting low abundant miRNAs. Using this assay, we have demonstrated that miRNAs are differentially expressed in the phloem sap and the surrounding vascular tissue. This method enables fast, sensitive and specific miRNA expression profiling and is suitable for facilitation of high-throughput detection and quantification of miRNA expression.