A Heat-Inducible Rubisco Activase Safeguards Photosynthesis via Multilevel Regulation and Dual Function

Rubisco activase (RCA) is critical for maintaining photosynthetic efficiency under heat stress, yet its regulatory mechanisms and functional diversification remain poorly understood. Here, we characterize a novel heat-inducible RCA family member (RCA1), which plays a pivotal role in mediating multidimensional thermotolerance in a tropical azalea. Both heterologous and homologous overexpression of RCA1 significantly mitigated heat-induced cellular damage and better maintained photosynthetic capacity, confirming its essential function in thermal resilience. Heat-induced RCA1 expression is transcriptionally activated by a synergistic HSFA1-HSFA2 complex and post-transcriptionally fine-tuned by extensive alternative splicing, forming a multilevel regulatory network under heat stress. Beyond its canonical Rubisco-activating enzymatic activity, RCA1 exhibits an additional thermoprotective function that suppresses the thermal aggregation of several chloroplast proteins, such as Lhca2, PetC and PsaK. RCA1 also interacts with the constitutive RCA2, likely through heteromeric RCA complex assembly, which helps retain RCA2 in a soluble pool under heat stress. Evolutionary analyses reveal functional divergence within the RCA family, with convergent evolution of heat-inducible RCA clades in Panicoideae grasses. Collectively, our findings uncover a novel dual-functional heat-inducible RCA, which provides a promising target for photosynthetic bioengineering to enhance crop thermotolerance and ensure agricultural sustainability under global warming.