ABSTRACT
Repulsive guidance molecule A (RGM A) was recently described as a potent inhibitor of neuroregeneration in a rat spinal cord injury model. The receptor mediating RGM A's repulsive activity was shown to be Neogenin, a member of the Deleted in Colorectal Cancer (DCC) family of netrin receptors. Binding of RGM A to Neogenin induces activation of the small GTPase RhoA and of its effector Rho-kinase by an unknown mechanism. Here we show, that the cytoplasmic tail of Neogenin interacts directly with the transcriptional coactivator LIM domain only 4 (LMO4) in human SH-SY5Y cells, human Ntera neurons, and in embryonic rat cortical neurons. RGM A binding to Neogenin but not binding of Netrin-1, induces release of LMO4 from Neogenin. Down-regulation of LMO4 neutralizes the repulsive activity of RGM A in neuronal cell lines and embryonic rat cortical neurons and prevents RhoA activation. These results show for the first time that an interaction of Neogenin with LMO4 is involved in the RGM A - Neogenin signal transduction pathway for RhoA activation.
Subject(s)
Homeodomain Proteins/metabolism , Membrane Glycoproteins/metabolism , Membrane Proteins/metabolism , Nerve Tissue Proteins/metabolism , Neurons/physiology , Transcription Factors/metabolism , Adaptor Proteins, Signal Transducing , Amides/pharmacology , Analysis of Variance , Animals , Cells, Cultured , Cerebral Cortex/cytology , Embryo, Mammalian , Enzyme Inhibitors/pharmacology , GPI-Linked Proteins , Gene Expression/drug effects , Green Fluorescent Proteins/biosynthesis , Homeodomain Proteins/biosynthesis , Humans , LIM Domain Proteins , Neurites/drug effects , Neurites/physiology , Neurons/cytology , Neurons/drug effects , Protein Structure, Tertiary , Pyridines/pharmacology , RNA, Small Interfering/pharmacology , Rats , Transcription Factors/biosynthesis , Transfection/methods , Tubulin/pharmacology , Two-Hybrid System Techniques , rhoA GTP-Binding Protein/genetics , rhoA GTP-Binding Protein/metabolismABSTRACT
Asthma is an increasing health problem worldwide, but the long-term temporal pattern of clinical symptoms is not understood and predicting asthma episodes is not generally possible. We analyse the time series of peak expiratory flows, a standard measurement of airway function that has been assessed twice daily in a large asthmatic population during a long-term crossover clinical trial. Here we introduce an approach to predict the risk of worsening airflow obstruction by calculating the conditional probability that, given the current airway condition, a severe obstruction will occur within 30 days. We find that, compared with a placebo, a regular long-acting bronchodilator (salmeterol) that is widely used to improve asthma control decreases the risk of airway obstruction. Unexpectedly, however, a regular short-acting beta2-agonist bronchodilator (albuterol) increases this risk. Furthermore, we find that the time series of peak expiratory flows show long-range correlations that change significantly with disease severity, approaching a random process with increased variability in the most severe cases. Using a nonlinear stochastic model, we show that both the increased variability and the loss of correlations augment the risk of unstable airway function. The characterization of fluctuations in airway function provides a quantitative basis for objective risk prediction of asthma episodes and for evaluating the effectiveness of therapy.
