ABSTRACT
In this study, we examined protein-protein interactions between two neuronal receptors, low density lipoprotein receptor-related protein (LRP) and sorLA/LR11, and found that these receptors interact, as indicated by three independent lines of evidence: co-immunoprecipitation experiments on mouse brain extracts and mouse neuronal cells, surface plasmon resonance analysis with purified human LRP and sorLA, and fluorescence lifetime imaging microscopy (FLIM) on rat primary cortical neurons. Immunocytochemistry experiments revealed widespread co-localization of LRP and sorLA within perinuclear compartments of rat primary neurons, while FLIM analysis showed that LRP-sorLA interactions take place within a subset of these compartments.
Subject(s)
LDL-Receptor Related Proteins/metabolism , Receptors, LDL/metabolism , Animals , Binding Sites , Cells, Cultured , Embryo, Mammalian , Green Fluorescent Proteins/genetics , Humans , Immunoprecipitation/methods , LDL-Receptor Related Proteins/genetics , Mice , Microscopy, Fluorescence , Neuroblastoma , Neurons/metabolism , Protein Binding , Protein Interaction Domains and Motifs , Protein Interaction Mapping/methods , Rats , Rats, Sprague-Dawley , Receptors, LDL/genetics , Surface Plasmon Resonance/methods , Transfection/methodsABSTRACT
Motor neuron (MN) mitochondrial abnormalities and elevation in spinal fluid levels of the inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) have been implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS). The mechanism of neuron death in ALS remains unclear, along with the contributions of mitochondrial dysfunction and inflammation in the process. Cell cultures enriched for MN derived from embryonic rat spinal cords were established and directly exposed in vitro to recombinant TNF-alpha for varying lengths of time. Although cytokine exposure for up to 4 days failed to induce MN death, mitochondrial changes were observed shortly after initiating treatment. Our results demonstrate that TNF-alpha induced mitochondrial redistribution toward the soma in MN. We postulate that inflammation may precede, and in fact cause, the mitochondrial changes observed in ALS tissue.