Yuanyuan Xu

Plant GATA transcription factors are zinc-finger proteins involved in signal transduction, growth and development. However, their potential involvement in plant height and gibberellin-associated shoot growth in pummelo insufficiently characterised. In this study, members of the CgGATA gene family were identified genome-wide and characterised through analyses of physicochemical properties, chromosomal distribution, phylogenetic relationships, gene structures, conserved motifs, collinearity and predicted cis-acting elements in putative upstream regulatory regions. RT-qPCR was used to examine expression profiles across sampled tissues, and expression changes associated with prolonged gibberellin (GA₃) treatment in ‘Wanbai Pummelo’ shoot tips, and expression patterns in tall and dwarf pummelo genotypes. A total of 23 CgGATA genes were identified and classified into four phylogenetic subfamilies. Members of the same subfamily generally shared similar gene structures and conserved-motif compositions. Comparative collinearity analysis identified more syntenic GATA gene pairs between pummelo and sweet orange than between pummelo and Arabidopsis thaliana. The putative upstream regulatory regions contained predicted elements associated with light, phytohormone, growth and stress responses. The genes exhibited distinct expression profiles across the sampled tissues, with many showing relatively high within-gene values in stems and leaves. Prolonged GA₃ treatment promoted seedling shoot elongation and was associated with changes in the transcript abundance of several CgGATA genes at 28 d. Integration of these expression profiles, the tall dwarf comparison and predicted gibberellin-responsive elements prioritised CgGATA11 and CgGATA18 as candidates for investigating plant height variation and shoot elongation in pummelo. These findings provide a basis for functional studies but do not establish that either gene directly regulates gibberellin signalling or plant height.

Understanding the molecular regulation of citric acid accumulation in citrus fruits is crucial, as acidity directly influences fruit flavor, consumer preference, and commercial value. Citric acid is the predominant organic acid in citrus, and its levels are shaped by several factors, including genetic and developmental factors. ‘Jinyan’ Bingtang orange (Citrus sinensis cv. Jinyan) is a novel mutant derived from ‘Jinhong’ Bingtang orange (C. sinensis cv. Jinhong) that has a noticeably sour taste. However, the molecular basis of the increased citrate content in ‘Jinyan’ fruits remains unclear. This study compared the organic acid profiles and expression of citric acid metabolism-related genes between ‘Jinyan’ and ‘Jinhong’ fruit juice sacs throughout fruit development. The trend of citric acid content in both cultivars was similar; however, ‘Jinyan’ consistently presented significantly higher levels than ‘Jinhong’ did from 95 to 215 days after flowering (DAF). After 155 DAF, the transcript levels of citrate biosynthesis-related genes (PEPC1, PEPC2, PEPC3, CS1, and CS2) and citrate transport-related genes (V₁-E1, V₁-E2, V₀-a2, V₀-d, VHP1, VHP2, and CsPH8) were significantly greater in ‘Jinyan’ than in ‘Jinhong’. In contrast, citrate degradation-related genes (NAD-IDH2 and NAD-IDH3) were expressed at lower levels than in ‘Jinhong’. Notably, the expression patterns of V₁-E2 and CsPH8 closely matched the changes in citrate content in both cultivars. These results indicate that, compared with ‘Jinhong’, high citric acid accumulation in the juice sacs of ‘Jinyan’ fruit is likely due to increased citrate synthesis (via upregulated PEPCs and CSs) and increased vacuolar citrate sequestration (via upregulated proton pumps and transporters), coupled with reduced citrate degradation (lower NAD-IDH2/3).

The citrus cultivar ‘Local Juhong’, which has historically been used as a traditional Chinese medicinal material, originated in Yuanjiang County, Hunan Province.Its parental type and genetic background are indistinct as of yet. Morphological observation shows that ‘Local Juhong’ has a slight oblateness in fruit shape, a relatively smooth pericarp, a fine and slightly raised oil vacuole, and an inward concave at the blossom end. The tree form and fruit and leaf morphology of ‘Local Juhong’ are similar to those of ‘Huangpi’ sour orange. To reveal the genetic background of ‘Local Juhong’, 21 citrus accessions were evaluated using nuclear and chloroplast SSR markers and whole-genome SNP information. ‘Local Juhong’ was grouped with mandarins and sub-grouped with ‘Miyagawa Wase’ and ‘Yanxi Wanlu’ in a nuclear SSR analysis, which indicated that its pollen parent might be mandarins. It was closely clustered with orange and pummelo in the chloroplast SSR analysis. The genomic sequence similarity rate of ‘Local Juhong’ with mandarin and pummelo heterozygosity was 70.88%; the main part was the heterozygosity, except for the unknown (19.66%), mandarin (8.73%), and pummelo (3.9%) parts. Thus, ‘Local Juhong’ may be an F₁ hybrid with pummelo as the female parent and mandarin as the male parent, sharing sisterhood with ‘Huangpi’ sour orange.

Although revisiting the discoveries and implications of genetic variations using phased genomics is critical, such efforts are still lacking. Somatic mutations represent a crucial source of genetic diversity for breeding and are especially remarkable in heterozygous perennial and asexual crops. In this study, we focused on a diploid sweet orange (Citrus sinensis) and constructed a haplotype-resolved genome using high fidelity (HiFi) reads, which revealed 10.6% new sequences. Based on the phased genome, we elucidate significant genetic admixtures and haplotype differences. We developed a somatic detection strategy that reveals hidden somatic mutations overlooked in a single reference genome. We generated a phased somatic variation map by combining high-depth whole-genome sequencing (WGS) data from 87 sweet orange somatic varieties. Notably, we found twice as many somatic mutations relative to a single reference genome. Using these hidden somatic mutations, we separated sweet oranges into seven major clades and provide insight into unprecedented genetic mosaicism and strong positive selection. Furthermore, these phased genomics data indicate that genomic heterozygous variations contribute to allele-specific expression during fruit development. By integrating allelic expression differences and somatic mutations, we identified a somatic mutation that induces increases in fruit size. Applications of phased genomics will lead to powerful approaches for discovering genetic variations and uncovering their effects in highly heterozygous plants. Our data provide insight into the hidden somatic mutation landscape in the sweet orange genome, which will facilitate citrus breeding.