Asha L. Bhakar

NRAGE, A Novel MAGE Protein, Interacts with the p75 Neurotrophin Receptor and Facilitates Nerve Growth Factor-Dependent Apoptosis

The mechanisms employed by the p75 neurotrophin receptor (p75NTR) to mediate neurotrophin-dependent apoptosis are poorly defined. Two-hybrid analyses were used to identify proteins involved in p75NTR apoptotic signaling, and a p75NTR binding partner termed NRAGE (for neurotrophin receptor-interacting MAGE homolog) was identified. NRAGE binds p75NTR in vitro and in vivo, and NRAGE associates with the plasma membrane when NGF is bound to p75NTR. NRAGE blocks the physical association of p75NTR with TrkA, and, conversely, TrkA overexpression eliminates NRAGE-mediated NGF-dependent death, indicating that interactions of NRAGE or TrkA with p75NTR are functionally and physically exclusive. NRAGE overexpression facilitates cell cycle arrest and permits NGF-dependent apoptosis within sympathetic neuron precursors cells. Our results show that NRAGE contributes to p75NTR-dependent cell death and suggest novel functions for MAGE family proteins.

Nuclear factor kB signaling regulates neuronal morphology and cocaine reward

Although chronic cocaine-induced changes in dendritic spines on nucleus accumbens (NAc) neurons have been correlated with behavioral sensitization, the molecular pathways governing these structural changes, and their resulting behavioral effects, are poorly understood. The transcription factor, nuclear factor κ B (NFκB), is rapidly activated by diverse stimuli and regulates expression of many genes known to maintain cell structure. Therefore, we evaluated the role of NFκB in regulating cocaine-induced dendritic spine changes on medium spiny neurons of the NAc and the rewarding effects of cocaine. We show that chronic cocaine induces NFκB-dependent transcription in the NAc of NFκB-Lac transgenic mice. This induction of NFκB activity is accompanied by increased expression of several NFκB genes, the promoters of which show chromatin modifications after chronic cocaine exposure consistent with their transcriptional activation. To study the functional significance of this induction, we used viral-mediated gene transfer to express either a constitutively active or dominant-negative mutant of Inhibitor of κ B kinase (IKKca or IKKdn), which normally activates NFκB signaling, in the NAc. We found that activation of NFκB by IKKca increases the number of dendritic spines on NAc neurons, whereas inhibition of NFκB by IKKdn decreases basal dendritic spine number and blocks the increase in dendritic spines after chronic cocaine. Moreover, inhibition of NFκB blocks the rewarding effects of cocaine and the ability of previous cocaine exposure to increase an animals preference for cocaine. Together, these studies establish a direct role for NFκB pathways in the NAc to regulate structural and behavioral plasticity to cocaine.

The p75 Neurotrophin Receptor (p75NTR) Alters Tumor Necrosis Factor-mediated NF-κB Activity under Physiological Conditions, but Direct p75NTR-mediated NF-κB Activation Requires Cell Stress

The p75 neurotrophin receptor (p75NTR) has been linked to activation of the NF-κB transcriptional complex in oligodendrocytes, Schwann cells, and PCNA cells. In this report, tumor necrosis factor (TNF)- and neurotrophin-mediated NF (nuclear factor)-κB activation were compared in several cell lines. All cell types showed TNF-mediated activation of NF-κB, but direct neurotrophin-dependent activation of NF-κB was never observed under normal growth conditions. In PCNA cells, a modest nerve growth factor (NGF)-dependent induction of NF-κB was detected but only after cells were subjected to severe stress. Although NGF binding did not directly activate NF-κB under normal conditions, NGF consistently altered TNF-dependent NF-κB activation in each cell type examined, and extended exposure to NGF and TNF always increased NF-κB activation over that achieved with TNF alone. The increase in NF-κB activity mediated by NGF correlated with reduced levels of IκBα; NGF added alone had no effect on IκBα levels, but when added with TNF, NGF treatment significantly reduced IκBα levels. We propose that modulation of cytokine receptor signaling is a significant physiological function of the p75 neurotrophin receptor and that previous reports of direct NF-κB activation through p75NTR reflect this modulatory activity.

TRAF6‐dependent NF‐kB transcriptional activity during mouse development

Nuclear factor‐kappa B (NF‐kB) transcriptional activity is induced by numerous stimuli. To identify tissues exhibiting NF‐kB transcriptional activity during development, we analyzed transgenic reporter mice that express β‐galactosidase from an NF‐kB–responsive element. We report that NF‐kB activation is widespread and present in numerous epithelial structures and within vasculature. Several regions of the developing central nervous system, including the roof plate and floor plate of the midbrain, show prominent NF‐kB activation. To assess the role of the TRAF6 adaptor protein in developmental NF‐kB activity, we analyzed NF‐kB activation in reporter mice rendered null for TRAF6. Deletion of TRAF6 resulted in the loss of NF‐kB activity in epithelia, in vasculature, and in roof and floor plate but had no effect on NF‐kB activity developing telencephalon, choroid plexus, cochlear canal, and thymus. These data indicate that NF‐kB transcriptional activity is present in a broad range of structures during development and that TRAF6 plays a critical role mediating developmental NF‐kB activation in many but not all tissues. Developmental Dynamics 231:122–127, 2004.