Functional Characterization of PpGS2 Promoter in Response to Hormones and Low Nitrogen in Kentucky Bluegrass

Kentucky bluegrass ( Poa pratensis L.) is an important cool-season turfgrass with high nitrogen (N) fertilizer requirements. PpGS2 is a key gene for nitrogen uptake assimilation, but the core regulatory region of its promoter remains unclear. To characterize the transcriptional regulation of the PpGS2 promoter, a series of 5’-terminal deletion constructs were generated, each fused to the β-glucuronidase (GUS) reporter gene and transformed into Arabidopsis and tobacco. All fragments exhibited transcriptional activity, with activity declining as fragment length decreased. The full-length promoter drove GUS expression most strongly in the leaves of transgenic Arabidopsis. In addition, exogenous hormones, low nitrogen, and light stress all significantly enhanced promoter activity, with methyl jasmonate (MeJA) showing the strongest induction. Deletion analysis further revealed the distribution of response elements: Gibberellin (GA)‑responsive elements were predominantly localized between − 2000 bp and − 1600 bp; Abscisic acid (ABA)‑ and light‑responsive elements were mainly concentrated within the − 1200 bp to -800 bp and − 1600 bp to -800 bp ranges, respectively; Auxin (IAA) and MeJA responses relied on the synergy of multiple regions; The low-nitrogen response was mediated by both positive and negative cis-acting elements. The − 2000bp to -1200 bp region likely contained inhibitory elements, whereas the − 1200bp to -500 bp region harbored enhancing elements. In conclusion, this study systematically elucidated the regulatory mode of the PpGS2 promoter for the first time, thereby enabling a better understanding of the synergistic regulatory mechanism of PpGS2 under hormonal and environmental signals.