Genome-wide expression analysis of soybean MADS genes showing potential function in the seed development

Abstract

The MADS family is an ancient and best-studied transcription factor and plays fundamental roles in almost every developmental process in plants. In the plant evolutionary history, the whole genome duplication (WGD) events are important not only to the plant species evolution, but to expansion of members of the gene families. Soybean as a model legume crop has experience three rounds of WGD events. Members of some MIKC(C) subfamilies, such as SOC, AGL6, SQUA, SVP, AGL17 and DEF/GLO, were expanded after soybean three rounds of WGD events. And some MIKC(C) subfamilies, MIKC* and type I MADS families had experienced faster birth-and-death evolution and their traces before the Glycine WGD event were not found. Transposed duplication played important roles in tandem arrangements among the members of different subfamilies. According to the expression profiles of type I and MIKC paralog pair genes, the fates of MIKC paralog gene pairs were subfunctionalization, and the fates of type I MADS paralog gene pairs were nonfunctionalization. 137 out of 163 MADS genes were close to 186 loci within 2 Mb genomic regions associated with seed-relative QTLs, among which 115 genes expressed during the seed development. Although MIKC(C) genes kept the important and conserved functions of the flower development, most MIKC(C) genes showed potentially essential roles in the seed development as well as the type I MADS.

Figure 1. The soybean MADS gene family.

The gene names of MIKC^c, MIKC*, Mα, Mβ and Mγ were abbreviated as M1 to M163 and were in orange, purple, blue, green and yellow, respectively, and short lines in corresponding color in the red blocks showed their locations in the soybean genome (Table S1). The short black lines in the green arcs showed the markers associated with non-seed QTLs and those in the red short blocks showed the markers associated with seed QTL (Table S4). The red blocks showed regions of QTLs relative to the seed traits according to the markers (Table S4). The light blue rainbows showed collinear relationships among the blocks containing MADS genes according to the MCScanX results (Table S3) and the red curves showed the paralogs. Twenty chromosomes (GM1-20) were in different colors and the size of the arc showed the size of chromosome (Mb). The figure was created through the software Circos (http://circos.ca/).

Figure 2. Schematic diagrams of motif organizations.

MIKC^C can be grouped into two type: a and b (Figure S2). The average length of members of each five families was as the length of a family. Motif 1 and 2 are equivalent to the MADS-box domain (PF00319), and motif 3 and motif 6 is the part of the I-domain and K-box domain (PF01486) for type II MADS proteins, respectively. Other motifs was unknown. The detailed information of soybean MADS motif organizations referred to Figure S2.

Figure 3. In silico expression profiles and the evolutional pattern of soybean MIKC genes.

The RNA-seq relative expression data of 17 tissues was used to re-construct expression patterns of MIKC genes. 3 samples from soybean seed compartments: GloE (Globular stage embryo proper), SCP (Early maturation seed coat parenchyma) and GloS (Globular stage suspensor); 10 soybean tissues samples: Gs (Globular Stage Seed), Hs (Heart Stage Seed), Cs (Cotyledon Stage Seed), Es (Early Maturation Stage Seed), Ds (Dry Seed), R (Root), S (Stem), L (Trifoliate leave), F (Floral bud), and WS (Whole seedling six days after imbibition); 4 soybean cotyledon development samples: CoM (Mid-maturation cotyledon), CoL (Late-maturation cotyledon), CoD (Dry seed) and CoS (Seedling cotyledon). The raw data was downloaded from the website http://seedgenenetwork.net/presentation. Gene names in red showed dispersed duplicate, in blue showed proximal duplicate, and in green its paralog genes were lost during evolution. The lines showed the blocks containing the corresponding MADS genes experienced the WGD events, and the evolution models of the blocks were displayed in Figure 1 and S5. The raw relative expressions of 163 MADS genes were in the Table S6.

Figure 4. In silico expression profiles and the evolutional pattern of soybean type I MADS genes.

Notes as Figure 3.

Figure 5. Expression cluster analysis based on in silico expression of 138 MADS genes.

The samples were designed as Figure 3. The gray line shows expression profiles of each genes, and the green line is the average expression and indicates the expression pattern of one cluster. For simplicity, gene names display the corresponding code numbers of every subfamily (Figure S1 and Table S6).

Figure 6. Expression patterns of GmMADS genes in the seeds by RT-qPCR.

4 samples at the seedling stage (the unifoliolates open fully): U-R (roots), U-S (stems), U-C (cotyledons) and U-U (leaves); 4 samples at the flowering stage: F-R (roots), F-S (stems), F-L (leaves) and F (flowers); 4 seed development samples: S1 (seeds at 7 days after flowering), S2 (seeds at 14 days after flowering), S3 (seeds at 21 days after flowering) and S4 (dry seeds). The similar expression profiles were in the similar color background. The bar is the average with standard deviation of the expression levels among three different replicates. The geometric means of GmSKIP16, GmUNKI and GmUNKII transcripts were used as the reference transcript. The values are means of three replicates, and each replicate represented a pool from at least five plants. Error bars represent SD.

Figure 7. Expression peaks in the flowers through RT-qPCR. Notes as Figure 6.

Figure 8. High expression in the seeds and flowers through RT-qPCR.

Notes as Figure 6.

Figure 9. High expression in the flowers and leaves through RT-qPCR.

Notes as Figure 6.

Figure 10. Expression peaks in the leaves through RT-qPCR.

Notes as Figure 6.

Figure 11. Expression peaks in the stems through RT-qPCR.

Notes as Figure 6.

Figure 12. Expression peaks in the roots through RT-qPCR.

Notes as Figure 6.

Figure 13. Expression divergence of paralog gene pairs.

The upper triangles showed the expression of the lift genes of the paralog gene pairs, and the lower triangle the expression of the right genes of the paralog gene pairs. The raw relative expressions of 163 MADS genes were in the Tab S6. Other notes were similar to Figure 3.