Genome-Wide Identification of the Transcription Factors Involved in Citrus Fruit Ripening from the Transcriptomes of a Late-Ripening Sweet Orange Mutant and Its Wild Type

Abstract

Fruit ripening is a genetically programmed process. Transcription factors (TFs) play key roles in plant development and ripening by temporarily and spatially regulating the transcription of their target genes. In this study, a total of 159 TFs were identified from a spontaneous late-ripening mutant 'Fengwan' (C. sinensis L. Osbeck) sweet orange (MT) and its wild-type counterpart ('Fengjie 72-1', WT) along the ripening period via the Transcription Factor Prediction of PlantTFDB 3.0. Fifty-two differentially expressed TFs were identified between MT and WT; 92 and 120 differentially expressed TFs were identified in WT and MT, respectively. The Venn diagram analysis showed that 16 differentially expressed TFs were identified between MT and WT and during the ripening of WT and MT. These TFs were primarily assigned to the families of C2H2, Dof, bHLH, ERF, MYB, NAC and LBD. Particularly, the number of TFs of the ERF family was the greatest between MT and WT. According to the results of the WGCNA analysis, a weighted correlation network analysis tool, several important TFs correlated to abscisic acid (ABA), citric acid, fructose, glucose and sucrose were identified, such as RD26, NTT, GATA7 and MYB21/62/77. Hierarchical cluster analysis and the expression analysis conducted at five fruit ripening stages further validated the pivotal TFs that potentially function during orange fruit development and ripening.

Fig 1. The Venn Diagram analysis (A) and the families assignment of TFs of WT (B), MT (C) and MT vs WT (D).

MT vs WT indicate the TF cluster, which is differentially expressed between MT and WT.

Fig 2. Biological process (A) and molecular function (B) enrichment analysis of the TFs differentially expressed during fruit ripening in both MT and WT.

Bubble color indicates the p-value; plot size indicates the frequency of the GO term in the underlying GOA database (bubbles of more general terms are larger).

Fig 3. Hierarchical cluster analysis of the TF differential expressed between MT and WT.

Fig 4. Hierarchical cluster tree with dissimilarity based on topological overlap showing coexpression modules identified by WGCNA (A).

Each leaf in the tree is one gene. The major tree branches constitute 32 modules labeled by different colors. Module colors were determined in the single-block analysis. B, Module-physiological traits association. Each row corresponds to a module. The number of genes in each module is indicated on the left. Each column corresponds to a physiological trait. The color of each cell at the row-column intersection indicates the correlation coefficient between the module and the physiological trait, and the numbers in each cell indicate correlation coefficient R and P value, respectively.

Fig 5. Module-TF association.

Each row corresponds to a module. The number of genes in each module is indicated on the left. Each column corresponds to a TF. The color of each cell at the row-column intersection indicates the correlation coefficient between the module and the TF, and the numbers in each cell indicate correlation coefficient R and P value, respectively.

Fig 6. Expression analysis of TFs at five citrus fruit ripening stages.

A, B and C indicate three expression patterns between MT and WT. 150, 170, 190, 210 and 240 indicate 150, 170, 190, 210 and 240 DAF, respectively. A single asterisk (*) represents a statistically significant difference (P < 0.05). Analyzed using Student's t-test.