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Abstract
LEAFY COTYLEDON1 (LEC1) is a HAP3 subunit of CCAAT-binding transcription factor, which controls several aspects of embryo and postembryo development, including embryo morphogenesis, storage reserve accumulation and skotomorphogenesis. Herein, using the method of chromosomal walking, a 2707bp upstream sequence from the ATG initiation codon site of AhLEC1A which is a homolog of Arabidopsis LEC1 was isolated in peanut. Its transcriptional start site confirmed by 5' RACE was located at 82 nt from 5' upstream of ATG. The bioinformatics analysis revealed that there existed many tissue-specific elements and light responsive motifs in its promoter. To identify the functional region of the AhLEC1A promoter, seven plant expression vectors expressing the GUS (β-glucuronidase) gene, driven by 5' terminal series deleted fragments of AhLEC1A promoter, were constructed and transformed into Arabidopsis. Results of GUS histochemical staining showed that the regulatory region containing 82bp of 5' UTR and 2228bp promoter could facilitate GUS to express preferentially in the embryos at different development periods of Arabidopsis. Taken together, it was inferred that the expression of AhLEC1A during seed development of peanut might be controlled positively by several seed-specific regulatory elements, as well as negatively by some other regulatory elements inhibiting its expression in other organs. Moreover, the GUS expression pattern of transgenic seedlings in darkness and in light was relevant to the light-responsive elements scattered in AhLEC1A promoter segment, implying that these light-responsive elements harbored in the AhLEC1A promoter regulate skotomorphogenesis of peanut seeds, and AhLEC1A expression was inhibited after the germinated seedlings were transferred from darkness to light.
Highlights
Several cis-elements in AhLEC1A promoter, such as SKn-1, CANBANAPA (CA)n, AGL15, DOF core, SEF3 motif and the like, which previously were demonstrated to be required for seed development and storage accumulation, were identified
Skn-1 motif and (CA)n element were reported to play a vital role in determining the seed-specific expression; the deletion of Skn-1 motif or (CA)n element in glutelin and napin promoter decreased their transcription in seeds [33, 34]
SEF3 motif is the binding site of Soybean Embryo Factor 3, which regulate the transcription of the β-conglycinin gene and participate in seed development [37, 38]
Summary
Seed development is a complex procedure of the flowering plant in life cycle, which can conceptually be divided into two distinct phases: embryo morphogenesis and seed maturation. Cloning and functional characterization of AhLEC1A promoter the Major Basic Research Project of Natural Science Foundation of Shandong Province (2018GHZ007). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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