Identification of Malus sieversii ABA receptor PYL8 interacting proteome using Y2H-seq.

The seed dormancy of Malus sieversii belongs to the comprehensive type of dormancy, abscisic acid (ABA) is one of the important hormones to lift the seed dormancy. The ABA receptor pyrabactin resistance-like (PYL) protein is involved in seed germination and various stress responses. Fourteen MsPYL genes were evaluated in the genome of Malus sieversii. Phylogenetic analyses demonstrated that MsPYL proteins can be divided into three groups. The promoter regions of MsPYL genes contain cis-acting elements associated with expression induction by abiotic stresses such as abscisic acid, drought, and low temperature. Arabidopsis transgenic lines overexpressing MsPYL8 exhibited a markedly reduced germination rate, an extended growth period, and a reduction in the biomass of the aboveground parts compared to the wild type. Additionally, we found that MsPYL8 interacted with 30 proteins, including low-temperature-induced 65 kDa protein-like (LTI) and late embryogenesis abundant (LEA). The results demonstrated that PYL8 binds to ABA during seed germination, inhibits the dephosphorylation activity of protein phosphatases type 2C (PP2C), and activates the transmission of sucrose non-fermenting-1-related protein kinases 2s (SnRK2s) to abscisic acid insensitive 5 (ABI5) signaling, thereby activating the expression of ABA-responsive genes to regulate ABA signal transduction. The screening and validation of PYL8 interaction with LTI, and LTI interaction with ABI5, provides a foundation for further elucidation of the mechanism of PYL gene involvement in Malus sieversii seed dormancy lifting under the regulation of the ABA signaling pathway.