N-Acetylcysteine Administration Attenuates Sensorimotor Impairments Following Neonatal Hypoxic-Ischemic Brain Injury in Rats.

Hypoxic ischemic (HI) brain injury that occurs during neonatal period has been correlated with severe neuronal damage, behavioral deficits and infant mortality. Previous evidence indicates that N-acetylcysteine (NAC), a compound with antioxidant action, exerts a potential neuroprotective effect in various neurological disorders including injury induced by brain ischemia. The aim of the present study was to investigate the role of NAC as a potential therapeutic agent in a rat model of neonatal HI brain injury and explore its long-term behavioral effects. To this end, NAC (50 mg/kg/dose, i.p.) was administered prior to and instantly after HI, in order to evaluate hippocampal and cerebral cortex damage as well as long-term functional outcome. Immunohistochemistry was used to detect inducible nitric oxide synthase (iNOS) expression. The results revealed that NAC significantly alleviated sensorimotor deficits and this effect was maintained up to adulthood. These improvements in functional outcome were associated with a significant decrease in the severity of brain damage. Moreover, NAC decreased the short-term expression of iNOS, a finding implying that iNOS activity may be suppressed and that through this action NAC may exert its therapeutic action against neonatal HI brain injury.

A cell-based chemical-genetic screen for amino acid stress response inhibitors reveals torins reverse stress kinase GCN2 signaling.

mTORC1 and GCN2 are serine/threonine kinases that control how cells adapt to amino acid availability. mTORC1 responds to amino acids to promote translation and cell growth while GCN2 senses limiting amino acids to hinder translation via eIF2α phosphorylation. GCN2 is an appealing target for cancer therapies because malignant cells can harness the GCN2 pathway to temper the rate of translation during rapid amino acid consumption. To isolate new GCN2 inhibitors, we created cell-based, amino acid limitation reporters via genetic manipulation of Ddit3 (encoding the transcription factor CHOP). CHOP is strongly induced by limiting amino acids and in this context, GCN2-dependent. Using leucine starvation as a model for essential amino acid sensing, we unexpectedly discovered ATP-competitive PI3 kinase-related kinase inhibitors, including ATR and mTOR inhibitors like torins, completely reversed GCN2 activation in a time-dependent way. Mechanistically, via inhibiting mTORC1-dependent translation, torins increased intracellular leucine, which was sufficient to reverse GCN2 activation and the downstream integrated stress response including stress-induced transcriptional factor ATF4 expression. Strikingly, we found that general translation inhibitors mirrored the effects of torins. Therefore, we propose that mTOR kinase inhibitors concurrently inhibit different branches of amino acid sensing by a dual mechanism involving direct inhibition of mTOR and indirect suppression of GCN2 that are connected by effects on the translation machinery. Collectively, our results highlight distinct ways of regulating GCN2 activity.

RHO to the DOCK for GDP disembarking: Structural insights into the DOCK GTPase nucleotide exchange factors.

The human dedicator of cytokinesis (DOCK) family consists of 11 structurally conserved proteins that serve as atypical RHO guanine nucleotide exchange factors (RHO GEFs). These regulatory proteins act as mediators in numerous cellular cascades that promote cytoskeletal remodeling, playing roles in various crucial processes such as differentiation, migration, polarization, and axon growth in neurons. At the molecular level, DOCK DHR2 domains facilitate nucleotide dissociation from small GTPases, a process that is otherwise too slow for rapid spatiotemporal control of cellular signaling. Here, we provide an overview of the biological and structural characteristics for the various DOCK proteins and describe how they differ from other RHO GEFs and between DOCK subfamilies. The expression of the family varies depending on cell or tissue type, and they are consequently implicated in a broad range of disease phenotypes, particularly in the brain. A growing body of available structural information reveals the mechanism by which the catalytic DHR2 domain elicits nucleotide dissociation and also indicates strategies for the discovery and design of high-affinity small-molecule inhibitors. Such compounds could serve as chemical probes to interrogate the cellular function and provide starting points for drug discovery of this important class of enzymes.