Population-genetic analysis of HvABCG31 promoter sequence in wild barley (Hordeum vulgare ssp. spontaneum)

Abstract

Background: The cuticle is an important adaptive structure whose origin played a crucial role in the transition of plants from aqueous to terrestrial conditions. HvABCG31/Eibi1 is an ABCG transporter gene, involved in cuticle formation that was recently identified in wild barley (Hordeum vulgare ssp. spontaneum). To study the genetic variation of HvABCG31 in different habitats, its 2 kb promoter region was sequenced from 112 wild barley accessions collected from five natural populations from southern and northern Israel. The sites included three mesic and two xeric habitats, and differed in annual rainfall, soil type, and soil water capacity.

Results: Phylogenetic analysis of the aligned HvABCG31 promoter sequences clustered the majority of accessions (69 out of 71) from the three northern mesic populations into one cluster, while all 21 accessions from the Dead Sea area, a xeric southern population, and two isolated accessions (one from a xeric population at Mitzpe Ramon and one from the xeric 'African Slope' of "Evolution Canyon") formed the second cluster. The southern arid populations included six haplotypes, but they differed from the consensus sequence at a large number of positions, while the northern mesic populations included 15 haplotypes that were, on average, more similar to the consensus sequence. Most of the haplotypes (20 of 22) were unique to a population. Interestingly, higher genetic variation occurred within populations (54.2%) than among populations (45.8%). Analysis of the promoter region detected a large number of transcription factor binding sites: 121-128 and 121-134 sites in the two southern arid populations, and 123-128,125-128, and 123-125 sites in the three northern mesic populations. Three types of TFBSs were significantly enriched: those related to GA (gibberellin), Dof (DNA binding with one finger), and light.

Conclusions: Drought stress and adaptive natural selection may have been important determinants in the observed sequence variation of HvABCG31 promoter. Abiotic stresses may be involved in the HvABCG31 gene transcription regulations, generating more protective cuticles in plants under stresses.

Figure 1

Haplotype distribution in five natural populations of Hordeum vulgare ssp. s (Spontaneum) in Israel. The white sectors in the pies indicate the frequencies of haplotypes unique to that population, while patterned sectors denote the frequencies of haplotypes shared between populations (for frequency details, see Table 2). P1, Mitzpe Ramon; P2, Dead Sea; P3, "Evolution Canyon"; P4, Arbel; P5, Yehudiyya.

Figure 2

Maximum likelihood bootstrap consensus phylogeny of 2 kb sequence of the HvABCG31 promoter. The phylogenetic consensus tree was constructed using the method of maximum likelihood in MEGA5. Values at the nodes are bootstrap percentage > 60%. Percentages below the cluster names refer to the accession proportions in specific populations. P1, Mitzpe Ramon; P2, Dead Sea; P3, "Evolution Canyon"; P4, Arbel; P5, Yehudiyya.

Figure 3

Differences between haplotypes and the consensus sequence of HvABCG31 promoter in Hordeum vulgare ssp. spontaneum. The number of rare polymorphic sites for each haplotype are above the bars. P1, Mitzpe Ramon; P2, Dead Sea; P3, "Evolution Canyon"; P4, Arbel; P5, Yehudiyya.

Figure 4

Numbers and categories of transcription factor binding sites (TFBSs) in the HvABCG31 promoter of Hordeum vulgare ssp. spontaneum. Different grids in the histogram refer to specific transcription factor binding sites; Sites indicated in red occurred at significantly higher frequencies than random, based on a permutation test with 10,000 runs; their sequences are given and related transcription factors are listed in brackets.

Figure 5

Proportions of conserved and non-conserved TFBSs in the Hordeum vulgare ssp. spontaneum HvABCG31 promoter sequence. The TFBSs were analysed using PlantPan database ( http://plantpan.mbc.nctu.edu.tw/index.php). The upper pie chart showed the over all proportions of conserved and non-conserved TFBSs, and the two lower pie charts showed the proportions of different categories of TFBSs within each group.

Figure 6

Distribution of non-conserved TFBSs in all 22 haplotypes of the HvABCG31 promoter. The upper dashed area refer to TFBSs that occurred in a majority of haplotypes, and the lower dashed area refer to those occurring in a minority of haplotypes.