Postnatal deamidation of 4E-BP2 in brain enhances its association with raptor and alters kinetics of excitatory synaptic transmission.

The eIF4E-binding proteins (4E-BPs) repress translation initiation by preventing eIF4F complex formation. Of the three mammalian 4E-BPs, only 4E-BP2 is enriched in the mammalian brain and plays an important role in synaptic plasticity and learning and memory formation. Here we describe asparagine deamidation as a brain-specific posttranslational modification of 4E-BP2. Deamidation is the spontaneous conversion of asparagines to aspartates. Two deamidation sites were mapped to an asparagine-rich sequence unique to 4E-BP2. Deamidated 4E-BP2 exhibits increased binding to the mammalian target of rapamycin (mTOR)-binding protein raptor, which effects its reduced association with eIF4E. 4E-BP2 deamidation occurs during postnatal development, concomitant with the attenuation of the activity of the PI3K-Akt-mTOR signaling pathway. Expression of deamidated 4E-BP2 in 4E-BP2(-/-) neurons yielded mEPSCs exhibiting increased charge transfer with slower rise and decay kinetics relative to the wild-type form. 4E-BP2 deamidation may represent a compensatory mechanism for the developmental reduction of PI3K-Akt-mTOR signaling.

Combinations of DEAD box proteins distinguish distinct types of RNA: protein complexes in neurons.

Transport of mRNAs to axons and dendrites in neurons is important for growth, polarization and plasticity. Recent proteomic studies in neurons have identified a number of DEAD box proteins as components of RNA granules. Using DEAD box proteins as markers, we have defined classes of RNA:protein structures present in neurons. In particular, we demonstrate that the conjunction of DEAD box 1 and DEAD box 3 identifies a motile ribosome-containing RNA granule present in both axons and dendrites that is similar to the biochemically isolated RNA granule. Conjunction of DEAD box 1 and the novel protein CGI-99 defines a distinct complex in neurons. Attempts to define a P-body like structure with expression of DEAD box 6 and decapping enzymes suggest that this structure may be more complex in neuronal processes than in other compartments. These studies hint at a great complexity in RNA transport and storage in neuronal processes.

The nuclear receptor NR4A1 is regulated by SUMO modification to induce autophagic cell death.

NR4A is a nuclear receptor protein family whose members act as sensors of cellular environment and regulate multiple processes such as metabolism, proliferation, migration, apoptosis, and autophagy. Since the ligand binding domains of these receptors have no cavity for ligand interaction, their function is most likely regulated by protein abundance and post-translational modifications. In particular, NR4A1 is regulated by protein abundance, phosphorylation, and subcellular distribution (nuclear-cytoplasmic translocation), and acts both as a transcription factor and as a regulator of other interacting proteins. SUMOylation is a post-translational modification that can affect protein stability, transcriptional activity, alter protein-protein interactions and modify intracellular localization of target proteins. In the present study we evaluated the role of SUMOylation as a posttranslational modification that can regulate the activity of NR4A1 to induce autophagy-dependent cell death. We focused on a model potentially relevant for neuronal cell death and demonstrated that NR4A1 needs to be SUMOylated to induce autophagic cell death. We observed that a triple mutant in SUMOylation sites has reduced SUMOylation, increased transcriptional activity, altered intracellular distribution, and more importantly, its ability to induce autophagic cell death is impaired.