Dynamics and Endocytosis of Flot1 in Arabidopsis Require CPI1 Function
Yangyang Cao, Qizouhong He, Zengxing Qi, Yan Zhang, Liang Lu, Jingyuan Xue, Junling Li, Ruili Li
Abstract
Membrane microdomains are nano-scale domains (10–200 nm) enriched in sterols and sphingolipids. They have many important biological functions, including vesicle transport, endocytosis, and pathogen invasion. A previous study reported that the membrane microdomain-associated protein Flotillin1 (Flot1) was involved in plant development in Arabidopsis thaliana; however, whether sterols affect the plant immunity conveyed by Flot1 is unknown. Here, we showed that the root length in sterol-deficient cyclopropylsterol isomerase 1 (cpi1-1) mutants expressing Flot1 was significantly shorter than in control seedlings. The cotyledon epidermal cells in cpi1-1 mutants expressing Flot1 were smaller than in controls. Moreover, variable-angle total internal reflection fluorescence microscopy (VA-TIRFM) and single-particle tracking (SPT) analysis demonstrated that the long-distance Flot1-GFP movement was decreased significantly in cpi1-1 mutants compared with the control seedlings. Meanwhile, the value of the diffusion coefficient Ĝ was dramatically decreased in cpi1-1 mutants after flagelin22 (flg22) treatment compared with the control seedlings, indicating that sterols affect the lateral mobility of Flot1-GFP within the plasma membrane. Importantly, using confocal microscopy, we determined that the endocytosis of Flot1-GFP was decreased in cpi1-1 mutants, which was confirmed by fluorescence cross spectroscopy (FCS) analysis. Hence, these results demonstrate that sterol composition plays a critical role in the plant defense responses of Flot1.