ABSTRACT
Heregulins (HRG) are known as soluble secreted growth factors that, on binding and activating ErbB3 and ErbB4 cell surface receptors, are involved in cell proliferation, metastasis, survival, and differentiation in normal and malignant tissues. Previous studies have shown that some HRG1 splice variants are translocated to the nucleus. By investigating the subcellular localization of HRGalpha(1-241), nuclear translocation and accumulation in nuclear dot-like structures was shown in breast cancer cells. This subcellular distribution pattern depends on the presence of at least one of two nuclear localization sequences and on two domains on the HRG construct that were found to be necessary for nuclear dot formation. Focusing on the nuclear function of HRG, a mammary gland cDNA library was screened with the mature form of HRGalpha in a yeast two-hybrid system, and coimmunoprecipitation of endogenous HRG was done. The data reveal positive interactions of HRGalpha(1-241) with nuclear factors implicated in different biological functions, including transcriptional control as exemplified by interaction with the transcriptional repressor histone deacetylase 2. In addition, HRGalpha(1-241) showed transcriptional repression activity in a reporter gene assay. Furthermore, a potential of HRG proteins to form homodimers was reported and the HRG sequence responsible for dimerization was identified. These observations strongly support the notion that HRG1 splice variants have multifunctional properties, including previously unknown regulatory functions within the nucleus that are different from the activation of ErbB receptor signaling.
Subject(s)
Cell Physiological Phenomena , Neuregulin-1/metabolism , Receptor, ErbB-2/metabolism , Active Transport, Cell Nucleus , Animals , COS Cells , Cell Cycle Proteins/metabolism , Cell Line, Tumor , Cell Nucleus/metabolism , Chlorocebus aethiops , Cullin Proteins/metabolism , Dimerization , Down-Regulation/genetics , Epidermal Growth Factor/chemistry , Exons/genetics , Histone Deacetylase 2 , Histone Deacetylases/metabolism , Humans , Neuregulin-1/chemistry , Nuclear Proteins/metabolism , Protein Binding , Protein Structure, Tertiary , Repressor Proteins/metabolism , Sequence DeletionABSTRACT
Friedreich Ataxia (FRDA), the most prevalent of the inherited ataxias, is a multi-systemic disease with loss of sensory neurons and life-threatening hypertrophic cardiomyopathy as its most severe manifestations. Reduced levels of the mitochondrial protein frataxin lead to cell-damaging oxidative stress and consequently FRDA is considered as a model for more common neurodegenerative disorders in which reactive radicals and oxidative stress are involved. We have developed a cellular assay system that discriminates between fibroblasts from FRDA patients and unaffected donors on the basis of their sensitivity to pharmacological inhibition of de novo synthesis of glutathione. With this assay we observed that supplementation with selenium effectively improved the viability of FRDA fibroblasts, indicating that basal selenium concentrations are not sufficient to allow an adequate increase in the activity of certain detoxification enzymes (such as GPX). Furthermore, we characterized potential drug candidates and found that idebenone, a mitochondrially localized antioxidant that ameliorates cardiomyopathy in FRDA patients, as well as other lipophilic antioxidants protected FRDA cells from cell death. Our results also demonstrate for the first time that small-molecule GPX mimetics have potential as a novel treatment strategy for Friedreich Ataxia and presumably also for other neurodegenerative diseases with mitochondrial impairment.
