New insights into the origin of the B genome of hexaploid wheat: evolutionary relationships at the SPA genomic region with the S genome of the diploid relative Aegilops speltoides

Abstract

Background: Several studies suggested that the diploid ancestor of the B genome of tetraploid and hexaploid wheat species belongs to the Sitopsis section, having Aegilops speltoides (SS, 2n = 14) as the closest identified relative. However molecular relationships based on genomic sequence comparison, including both coding and non-coding DNA, have never been investigated. In an attempt to clarify these relationships, we compared, in this study, sequences of the Storage Protein Activator (SPA) locus region of the S genome of Ae. speltoides (2n = 14) to that of the A, B and D genomes co-resident in the hexaploid wheat species (Triticum aestivum, AABBDD, 2n = 42).

Results: Four BAC clones, spanning the SPA locus of respectively the A, B, D and S genomes, were isolated and sequenced. Orthologous genomic regions were identified as delimited by shared non-transposable elements and non-coding sequences surrounding the SPA gene and correspond to 35,268, 22,739, 43,397 and 53,919 bp for the A, B, D and S genomes, respectively. Sequence length discrepancies within and outside the SPA orthologous regions are the result of non-shared transposable elements (TE) insertions, all of which inserted after the progenitors of the four genomes divergence.

Conclusion: On the basis of conserved sequence length as well as identity of the shared non-TE regions and the SPA coding sequence, Ae speltoides appears to be more evolutionary related to the B genome of T. aestivum than the A and D genomes. However, the differential insertions of TEs, none of which are conserved between the two genomes led to the conclusion that the S genome of Ae. speltoides has diverged very early from the progenitor of the B genome which remains to be identified.

Figure 1

Identification of the 'SPA orthologous region' and comparative annotation of the homoeologous A, B, D and S sequences. (A) Scaled diagram of annotation results of the SPA locus region in which (CDS) (light blue), class I TEs (yellow blocks), class II TEs (green blocks), unclassified elements (grey), MITEs (vertical black flags) are shown. The remaining white spaces correspond to unassigned DNA (no features of annotation). Grey blocks represent sequence conservation between the different genomes defining the 'SPA orthologous region'. Genes are numbered as follow: 1: Pseudo tubulin gene; 2: SPA; 3: Putative cortical cell-delineating gene; 4: Putative kinesin gene. Eight class I TE displaying complete LTR and TSD suitable for the estimation of the insertion dates are highlighted with red stars. (B) Multipipmaker alignment using the sequence of the SPA orthologous region of the S genome of Ae. speltoides as a matrix compared with the 3 other sequences available, i.e. T. aestivum gB (top), gA (center), gD (bottom). Coloured blocks show the percentage of sequence identity (> 90 in red; between 50 to 90% in green). The SPA gene is indicated as a blue box.

Figure 2

Comparison of the Ae. speltoides sequence with the A, B D genome sequence of T. aestivum. The dot plot was performed using the DOTTER program with default parameters between Ae. speltoides gS (horizontal) and the T. aestivum gA, -gB, -gD genome (vertical) sequences. Annotation features identified for these sequences are reported on the corresponding axes. Gene numbers and names as well as color codes for TEs and other DNA sequence classes are as in figure 1. Diagonals on the dot plot output that represent nucleotide conservation between the two analyzed sequences are highlighted with dotted blue circles. The loss of micro-colinearity corresponds to diagonal breaks. 'SPA orthologous region' defined as conserved sequences between Ae. speltoides gS and T. aestivum -gA, -gB, -gD sequences are mentioned with plain arrows on the four annotation features. SPA gene is shown with dotted arrows on the dot plot out put.

Figure 3

Phylogenic analysis of the SPA protein among plant species. 4 rice (RISBZ1-2-4-5 respectively AB053475, AB021736, AB053473, AB053474), 2 barley (BLZ1-2 respectively BLZ2, Y10834), 2 maize (O2-OHP1 respectively AJ491297, L00623), 1 sorghum (SBO2, X71636), 4 Arabidopsis thaliana (BZO2H1-4 respectively NM178959, NM122389, NM122760, NM115319), 1 Nicotiana tabacum (BRLZ, AY061648), 1 Petroselinum crispum (CPRF-2, X58577) and 4 wheat (SPA-gA, -gB, -gD, -gS, present analysis) sequences are involved in the tree. Parameters used to construct the tree are mentioned in the material and method section.

Figure 4

Evolutionary structure of the 'Ancestral SPA Locus'. From the 'Ancestral SPA Locus' of 16 598 bp, nested insertions of identified TE are shown for the four sequences (gA, gB, gD, gS). Graphical motifs used to materialize class I, class II unclassified TE as well as MITE are mentioned on the figure.