Figure 8. Uncaging PI(4,5)P₂ distinguishes mechanism of lipid-binding.

Two different roles of lipid-binding proteins can be distinguished by lipid uncaging: protein A (e.g. CAPS) binds to PI(4,5)P₂ in order to bring it to the vesicle and fusion machinery (localization role); protein B (e.g. synaptotagmin-1) binds to PI(4,5)P₂ as an obligatory part of its mechanism (stoichiometric role). (a) In the control situation, protein A colocalizes PI(4,5)P₂ with protein B, leading to fusion. (b). Upon deletion of protein A, protein B is unable to interact with PI(4,5)P₂. Uncaging uncovers PI(4,5)P₂ underneath the vesicle, allowing protein B to interact and support secretion. Thus, the defect (lack of protein A) is bypassed by PI(4,5)P₂ uncaging. (c) Upon deletion of protein B, protein A still co-localizes PI(4,5)P₂ with the vesicle, but protein B is missing and PI(4,5)P₂ uncaging does not overcome the defect.