Mass spectrometry-based profiling of phosphoinositide: advances, challenges, and future directions

Phosphoinositides (PIPs), the phosphorylated derivatives of phosphatidylinositol (PI), are low-abundance yet critical components of eukaryotic membranes. They play pivotal roles in a wide array of cellular processes, including signal transduction, membrane trafficking, and cell motility. The seven PIP subclasses, generated by phosphorylation at the 3-, 4-, and 5-positions of the inositol ring, are tightly regulated in both spatial and temporal contexts. Dysregulation of PIP metabolism is associated with a range of diseases, including cancer, myopathy, and neurodegenerative and developmental disorders. While the importance of phosphorylation of the inositol ring is well established, recent studies have clarified the role of the fatty acyl chain composition of PIPs. This has resulted in a growing interest in analytical techniques that can determine fatty acyl chain profiles of PIPs. Over the past three decades, substantial advances have been made in mass spectrometry-based techniques, enabling detailed characterization of PIP molecular species, including their phosphate regioisomers. This review provides an overview of the development of mass spectrometric methods for analyzing PIPs, with a particular focus on those enabling the separation of PIP regioisomers and the profiling of their acyl chain composition.