Transcriptome Shock in Developing Embryos of a Brassica napus and Brassica rapa Hybrid

Abstract

Interspecific crosses that fuse the genomes of two different species may result in overall gene expression changes in the hybrid progeny, called 'transcriptome shock'. To better understand the expression pattern after genome merging during the early stages of allopolyploid formation, we performed RNA sequencing analysis on developing embryos of Brassica rapa, B. napus, and their synthesized allotriploid hybrids. Here, we show that the transcriptome shock occurs in the developing seeds of the hybrids. Of the homoeologous gene pairs, 17.1% exhibit expression bias, with an overall expression bias toward B. rapa. The expression level dominance also biases toward B. rapa, mainly induced by the expression change in homoeologous genes from B. napus. Functional enrichment analysis revealed significant differences in differentially expressed genes (DEGs) related to photosynthesis, hormone synthesis, and other pathways. Further study showed that significant changes in the expression levels of the key transcription factors (TFs) could regulate the overall interaction network in the developing embryo, which might be an essential cause of phenotype change. In conclusion, the present results have revealed the global changes in gene expression patterns in developing seeds of the hybrid between B. rapa and B. napus, and provided novel insights into the occurrence of transcriptome shock for harnessing heterosis.

Keywords: RNA sequencing; expression level dominance; homoeolog expression bias; transcriptome shock.

Figure 1

Phenotypes of the parents’ and their hybrids’ seed, and the synteny analysis between B. napus and B. rapa. (A) The seed phenotype comparisons of the hybrid and its parents within 30 days after pollination. Scale bars, 0.5 cm. (B) Seed numbers per silique and seed diameter 30 days after pollination. Error bars, standard deviation. (C) Dot plot of pairwise synteny between the B. napus and B. rapa genomes. A01–A10 on the vertical axis indicated the chromosomes of the AA genome. A01–C09 on the horizontal axis showed the chromosomes of the AACC genome.

Figure 2

Global analysis of transcriptome data. (A–D) Venn diagrams of the parents and F1 hybrid with gene expression 0 ≤ FPKM < 1, 1 ≤ FPKM < 10, 10 ≤ FPKM < 100, and FPKM ≥ 100, respectively. (E) Heatmap of the expression of 24,060 genes with FPKM ≥ 2 in all three materials using lg(FPKM + 1) as the metric. (F) Statistics of the expression patterns in AA, AAC, and AACC characterized by four sets of FPKM values. (G) Identification of DEGs in three materials. The ten genes with the most significant expression differences and related information were listed in Table S3. AAC vs. AACC up-regulation means that the gene is more expressed in AAC than in AACC.

Figure 3

Homoeolog expression bias analysis of homoeologous gene pairs in AAC. (A) Aⁿ-A^r genome homoeolog expression bias analyses. (B) Cⁿ-A^r genome homoeolog expression bias analyses. The number of homologous gene pairs, as well as their percentage, is indicated in the figure. The size of the circles is only used to distinguish differences in the gene expression of the parents. The ratio of the areas within the circles indicates differences in gene expression levels in hybrids.

Figure 4

Expression level dominance and relationships between the ELD and individual homoeolog expression levels. (A) ELD of the homologous gene pairs in A^r-Aⁿ and A^r-Cⁿ. (B) Relationships between ELD and individual homoeolog expression levels explained the phenomenon of ELD. The four columns in the figure correspond to the four ELD types of XI, IV, and IX columns corresponding to Figure 4A.

Figure 5

Analysis of the transcription factors in non-additive genes. (A) Transcription factor type statistics of A^r-ELD genes. (B) KEGG enrichment analysis of transcription factors. The size of the circle represents the number of genes enriched to this term, and the circle’s color represents the p-value. (C) Expression profiles of the transcription factors of A^r-ELD genes (FPKM > 10, log2 fold change >4) in the parents and hybrids.

Figure 6

A^r-ELD transcription factor interactions and function analysis. Blue circles represent target transcription factors. The yellow box represents the comment function terms. Gray lines represent the interactions. Colored lines indicate the functions of the protein.