Interacting proteome of PtDAL10 and PtDAL1 in Pinus tabuliformis

The optimal timing of the transition from vegetative growth to reproductive growth is critical for plant reproductive success, and the underlying regulatory mechanisms have been well studied in angiosperm model species, but relatively little in gymnosperms. Recently, DAL1, a MADS domain transcription factor (TF) gene that shows a conserved age-related expression profile in conifers, was found to serve as an age timer. However, how the DAL1 mediates the onset of reproductive growth remains poorly understood. Here, we have shown that the PtDAL1 directly regulates the PtDAL10 transcription by binding to its promoter region, resulting in a ternary MADS-domain TF complex with PtDAL10 and PtMADS11, two potential candidate regulators of the vegetative to reproductive transition in Chinese pine (Pinus tabuliformis). The PtDAL10 was progressively induced in new shoots with age and highly accumulated in male and female cones. Overexpression of PtDAL10 rescued the flowering of ft-10 and soc1-1-2 mutants in Arabidopsis. We provide insight into the molecular components associated with the PtDAL1, which integrates the vegetative to reproductive phase transition into age-mediated progressive development of the whole plant in conifers.

Methods
The Y2H-seq screening method was established and described as previously (Chen et al., 2021). The BD-PtDAL10 construct was used as bait and the yeast library as prey to screen for the interacting proteins. Each monoclonal colony was considered as an event. One thousand events were selected and sequenced by next generation sequencing (NGS). Sequencing was performed on the Illumina NovaSeq 6000 platform, which was provided by Beijing Genewiz Biotechnology Co (https://www.genewiz.com.cn/). The final non-redundant proteins interacting with PtDAL10 were identified.