Regulation of tapetum-specific <i>TA29</i> promoter by <i>cis</i> -elements binding positive and negative regulators

The tapetum-specific TA29 promoter from Nicotiana tabacum has been extensively applied for pollination control in hybrid seed production.Despite its widespread use, the elements regulating the tissue-specific activity have not been fully explored.This study conducted in silico and functional analyses to identify and characterize cis-elements within the TA29 Upstream Regulatory Module (URM).Motif discovery tools and a comparative analysis across tapetum/antherexpressed genes were used to identify conserved binding motifs; these corresponded to transcription factors NtMYB12, NtGT-1a, NtNAG1, and NtHD-ZIP.Mutagenesis of these motifs revealed their functional roles: NtMYB12, NtGT-1a, and NtNAG1 act as positive regulators, while NtHD-ZIP acts as a negative regulator of TA29 URM activity.The findings highlight the importance of combinatorial regulation involving both positive and negative transcription factors when modulating the spatial and temporal activity of the TA29 URM.