Nir1-Nir2 Heterodimerization Expands the Sensitivity and Dynamic Range of the Phosphatidylinositol Cycle

The phosphatidylinositol (PI) cycle sustains phosphoinositide-calcium signaling by replenishing plasma membrane phosphatidylinositol 4,5-bisphosphate (PIP2) consumed during receptor activation. Recruitment of the lipid transfer protein Nir2 to endoplasmic reticulum (ER)-plasma membrane (PM) contact sites (also known as ER-PM junctions) is central to this process, enabling transfer of PI from the ER to the PM for PIP2 resynthesis. Although the disease-associated Nir2 paralog Nir1 is essential for Nir2 recruitment under physiological conditions, the molecular basis of this regulation has remained unresolved. Here, we identify a conserved Nir Dimerization (NirD) domain in both Nir1 and Nir2 and determine the crystal structures. We show that NirD mediates preferential Nir1-Nir2 heterodimerization, which promotes Nir2 recruitment to ER-PM junctions and enhances PIP2 replenishment in stimulated cells. Furthermore, Nir1-Nir2 heterodimerization confers stimulus-strength-dependent, graded recruitment of Nir2, thereby broadening the sensitivity and dynamic range of PI cycle activity. Together, our findings uncover the structural mechanism underlying Nir1-dependent regulation of Nir2 and reveal paralog heterodimerization as a key strategy for scaling lipid transport at membrane contact sites to signaling demand, ensuring robust phosphoinositide homeostasis.