Nanoclusters of biosynthetic enzymes define the supramolecular organization of leukotriene synthesis on nuclear membranes

Abstract

Leukotrienes (LTs) initiate and amplify immune responses. The assembly of macromolecular complexes containing 5‐lipoxygenase (5‐LO) and 5‐lipoxygenase‐activating protein (FLAP) on nuclear membranes initiates LT synthesis. We combined fluorescence lifetime imaging microscopy (FLIM) to probe 5‐LO/FLAP interactions, with stochastic optical reconstruction microscopy (STORM) to delineate the distribution of 5‐LO and FLAP on the nuclear envelope. LT synthesis in RBL‐2H3 cells was triggered via the high‐affinity IgE receptor FcεRI. Though cells primed with IgE did not produce LTs, a fraction of 5‐LO translocated to nuclear membranes and interacted with FLAP. Crosslinking FcεRI with IgE and antigen initiated LT synthesis and enhanced 5‐LO translocation and its interaction with FLAP. 5‐LO formed very small clusters on nuclear membranes of IgE‐primed cells, but addition of antigen induced redistribution into larger nanoclusters. In control and IgE‐loaded cells, FLAP was uniformly distributed along nuclear membranes but reorganized into discrete clusters upon FcεRI crosslinking. Coordinated assembly of FLAP and 5‐LO into large supramolecular complexes accompanies and may control LT synthesis.This work was supported by T32 DK7540 (MT) and R01 AI068871 (MT, RJS).