Allelic variants of <i>IRKI</i> contribute to photosynthetic efficiency by regulating rubisco activase in <i>Populus</i>

Photosynthesis directly determines plant biomass accumulation by controlling carbon flow and energy input. Thus, increasing photosynthetic efficiency is a promising approach for boosting plant growth and yield. However, the genetic basis of photosynthesis in perennial woody plants remains largely unknown, and the causative alleles warrant comprehensive investigation. Here, we performed a genome-wide association study (GWAS) on photosynthetic traits in a natural population of Chinese white poplar (Populus tomentosa). We identified inflorescence and root apices receptor-like kinase-interacting protein (IRKI) as a causative gene of photosynthesis that is significantly associated with the activity of rubisco activase (Rca). The seventh leaves of PtoIRKI-overexpression (OE) plants exhibited a 27.77% increase in net photosynthetic rate (Pn), a 31.42% rise in starch content, and a 16.83% expansion in leaf area compared to wild-type plants, whereas ptoirki-knockdown (KD) plants displayed the opposite phenotypes. Further analyses indicated that PtoIRKI interacted with PtoRca to enhance Rca activity, leading to increases in the activation state of ribulose bisphosphate carboxylase oxygenase (rubisco) and photosynthetic efficiency. Importantly, we identified an elite haplotype, PtoIRKIhap2, which exhibited higher PtoIRKI expression and Pn than PtoIRKIhap1. Finally, we found that homeodomain-leucine zipper protein 1 (PtoHB1) specifically bound to the PtoIRKIhap2 promoter, thereby promoting PtoIRKI expression and photosynthetic efficiency, as validated by integrating machine learning models and molecular experiments. Our results shed light on the molecular mechanism through which PtoIRKI modulates photosynthetic efficiency. We also provide an excellent haplotype module, PtoHB1-PtoIRKIhap2-PtoRca, that can be used to improve the photosynthesis of woody plants via molecular breeding.