Comparative Analysis And Functional Study Of SQUAMOSA Promoter Binding Protein-like(SPL)genes In Wheat

Common wheat(Triticum aestivum) is an allohexaploid produced by spontaneous hybridizations between cultivated tetraploid wheat(Triticum turgidum, AABB) and diploid wheat(Aegilops tauschii, DD). It is one of the most important crops worldwide. Transcription factors play fundamental roles in the developmental processes of plants. SQUAMOSA promoter binding protein-like(SPL) genes encode a group of plant-specific transcription factors, with a variety of regulatory roles in higher plants. However, little work has been done about SPLs in wheat due to its large genome and many repetitive sequences. Using the draft genome sequences of T. urartu and Ae. tauschii, we analyzed the sequence, structure and phylogenetic relationship of SPL genes from diploid progenitors and hexaploid wheat. Meanwhile, we cloned TaSPL3 and TaSPL6 genes in hexaploid wheat, studying their gene structure and expression. The main results are as follows:1. Using the SBP domain of rice SPLs to BLAST, we found 19 SPL genes in T. urartu and Ae. tauschii respectively, in which there were 7 and 13 SPLs had complete SBP domain respectively. Phylogenetic analysis showed that SPL genes from the two speices were conservative in structures except SPL4, SPL5 and SPL8. Among them, 6 and 5 SPL genes respectively were found to contain miR156 target sites in T. urartu and Ae. tauschii, which may be regulated by miR156. By analysis of gene structure and promoter cis-element, we noticed that there was differentiation between SPL homologs from different diploid wheat. From the tissue-specific expression and stress analysis, we could see the similarity between homologs of SPL1, SPL3, SPL6, SPL12 and SPL15.2. Through the results from diploid wheat, we selected two candidate genes TaSPL3 and TaSPL6 for further study in hexaploid wheat Chinese Spring. We cloned the two genes, and used nulli-tetrosomic lines to map them on chromosomes 6 and chromosomes 5 respectively. Tissue-specific analysis indicated both TaSPL3 and TaSPL6 were most highly expressed in leaves. Sequences analysis and structure comparison showed that the A/B/D copies of TaSPL3 were highly homologous, but only A/B copies were conservative in TaSPL6.3. We found a duplicated segment of 47 bp in TaSPL6 D, which may lead to early translation termination and produced a new allele named TaSPL6 DR without SBP domain. Among Chinese wheat mini-core collection, we found 19 wheat varieties that contained TaSPL6 DR and seemed to be associated with wheat maturity and grain numbers per spike in Chinese landraces. Transcriptional activation assay suggested that TaSPL3 and TaSPL6 had transcriptional activation function, and transcriptional activation domain of TaSPL6 was located upstream of SBP domain. We also found the expression diversity between A/B/D copies of TaSPL6. TaSPL6 D could affect the expression of TaSPL6 A and TaSPL6 B. The expression of TaSPL6 D was reduced in wheat varieties containing TaSPL6 DR. Meanwhile, overexpression of TaSPL3 and TaSPL6 in Arabidopsis showed TaSPL3 could promote plant flowering which was similar to AtSPL3, while TaSPL6 could make leaves larger and may be implicated in biomass accumulation.In summary, we performed a comprehensive analysis of SPL genes in wheat. We also acquired the sequences, structure and phylogenetic information of SPL genes and cloned TaSPL3 and TaSPL6 in hexaploid wheat. Our work demonstrated that SPL genes play essential roles in wheat and are likely to help for future molecular breeding in wheat.