Identification of cis-acting elements recognized by transcription factor LlWOX11 in Lilium lancifolium

WOX transcription factors play important roles in plant developmental processes and mainly bind to the WOX-binding element to regulate gene expression. Previously, we characterized a WOX gene from Lilium lancifolium, LlWOX11, positively regulating bulbil formation, and showed that it bound to the motif of TTAATGAG. However, whether LlWOX11 could bind to other motifs is unclear. In this study, Transcription Factor-Centered Yeast One Hybrid (TF-Centered Y1H) was utilized to study the motifs recognized by LlWOX11, and five motifs with seven bases were obtained. In addition to three motifs containing known cis-acting elements: TCAACTC (CAREOSREP1), AGAAAGA (DOFCOREZM/POLLENILELAT52), ACAGTAT (CACTFTPPCA1), we identified that LlWOX11 could bind to two new motifs: TGCGAAA, TCCATCA. We further searched for the core sequences of these motifs by Y1H. Dual-luciferase assays (LUC), Electrophoretic mobility shift assays (EMSA) and chromatin immunoprecipitation (ChIP) were performed to further determine that these motifs were bound by LlWOX11 in the plant. In addition, we found that LlWOX11 inhibited the transcription of LlRR9 by binding to the screened motifs in the promoter and further promoted bulbil formation. These findings will help to further reveal the functions of the WOX protein and the molecular mechanism of bulbil formation regulated by LlWOX11.