Self-interaction pattern and targeted potential protein interaction networks of Arabidopsis CTP:phosphocholine cytidylyltransferase 1

Abstract

Phosphatidylcholine (PC) is a fundamental component of eukaryotic membranes, and its biosynthesis is tightly regulated to maintain membrane integrity and function. Despite the key role of CTP:phosphocholine cytidylyltransferase (CCT) in the rate-limiting step of PC synthesis, little is known about how CCT is modulated through protein-protein interactions (PPIs). In this study, we selected Arabidopsis thaliana CCT1 (AtCCT1) to investigate the potential regulatory network governing PC biosynthesis. Using yeast two-hybrid (Y2H) and bimolecular fluorescence complementation assays, we discovered that AtCCT1 forms self-association and interacts with its isoform AtCCT2. Importantly, AtCCT1 was also found to interact with importin α and β proteins, implying a potentially regulated transport mechanism. In addition, AtCCT1 and an Arabidopsis Sec14 family protein may also have interactions, which weakly activated reporter genes in the Y2H system but exhibited relatively stronger fluorescence in transformed tobacco leaf cells. Collectively, this study provides the first evidence of specific PPIs involving AtCCT1, offering new insight into the post-translational regulation of PC synthesis. These findings lay a foundation for future studies exploring how dynamic protein assemblies fine-tune membrane lipid metabolism, possibly in response to developmental or environmental conditions.