NGF causes TrkA to specifically attract microtubules to lipid rafts.

Membrane protein sorting is mediated by interactions between proteins and lipids. One mechanism that contributes to sorting involves patches of lipids, termed lipid rafts, which are different from their surroundings in lipid and protein composition. Although the nerve growth factor (NGF) receptors, TrkA and p75(NTR) collaborate with each other at the plasma membrane to bind NGF, these two receptors are endocytosed separately and activate different cellular responses.

High-resolution fractionation of signaling endosomes containing different receptors.

Receptor endocytosis is regulated by ligand binding, and receptors may signal after endocytosis in signaling endosomes. We hypothesized that signaling endosomes containing different types of receptors may be distinct from one another and have different physical characteristics. To test this hypothesis, we developed a high-resolution organelle fractionation method based on mass and density, optimized to resolve endosomes from other organelles.

Distinct signalling particles containing ERK/MEK and B-Raf in PC12 cells.

Although several multiprotein complexes containing MAPKs (mitogen-activated protein kinases) have been identified using overexpression of kinases and scaffold proteins, the components of the complexes and their physical properties at endogenous expression levels have not been defined. We characterized a large protein complex containing a nerve-growth-factor-activated ERK (extracellular-signal-regulated kinase) and MEK (MAPK/ERK kinase) in rat pheochromocytoma (PC12) cells. This protein complex fractionated into a high-speed pellet and was resistant to non-ionic detergent treatments that solubilized membranes.