In post‐mitotic neurons, the mechanisms of the apoptotic checkpoint that is activated by DNA damage remain unclear. Here we show that in cultured cortical neurons, the DNA damaging agent camptothecin (CPT) reduced transcription of rRNA and disrupted nucleolar staining for B23/nucleophosmin suggesting DNA damage‐induced nucleolar stress. Although CPT activated the pro‐apoptotic protein p53, the CPT‐induced nucleolar stress was unaffected by p53 inhibition. In addition, brain‐derived neurotrophic factor‐mediated protection from CPT‐induced apoptosis prevented neither nucleolar stress nor p53 activation. Therefore, inhibition of rRNA transcription might be upstream of the pro‐apoptotic p53 activity. Indeed, short hairpin RNA‐mediated inhibition of a RNA‐Polymerase‐I co‐factor, transcription initiation factor IA, attenuated rRNA transcription causing nucleolar stress and p53‐dependent neuronal apoptosis. The protein synthesis inhibitor cycloheximide blocked apoptosis that was induced by over‐expressed shTIF‐IA or active form of p53. Also, the general transcription inhibitor actinomycin D triggered nucleolar stress and activated p53. However, it did not induce apoptosis except at the low concentration of 0.05 μg/mL with stronger inhibitory activity against nucleolar than extranucleolar transcription. Hence, nucleolar stress‐activated apoptosis requires extranucleolar transcription. This study identifies the nucleoli of post‐mitotic neurons as sensors of DNA damage coupling reduced rRNA transcription to p53‐mediated apoptosis that requires de novo expression of protein‐coding genes. Thus, rDNA selectivity of DNA damage may determine its ability to induce neuronal apoptosis.

Chronic stress is a risk factor for the development of psychopathologies characterized by cognitive dysfunction and deregulated social behaviours. Emerging evidence suggests a role for cell adhesion molecules, including nectin-3, in the mechanisms that underlie the behavioural effects of stress. We tested the hypothesis that proteolytic processing of nectins by matrix metalloproteinases (MMPs), an enzyme family that degrades numerous substrates, including cell adhesion molecules, is involved in hippocampal effects induced by chronic restraint stress. A reduction in nectin-3 in the perisynaptic CA1, but not in the CA3, compartment is observed following chronic stress and is implicated in the effects of stress in social exploration, social recognition and a CA1-dependent cognitive task. Increased MMP-9-related gelatinase activity, involving N-methyl-D-aspartate receptor, is specifically found in the CA1 and involved in nectin-3 cleavage and chronic stress-induced social and cognitive alterations. Thus, MMP-9 proteolytic processing emerges as an important mediator of stress effects in brain function and behaviour.

Serum response factor (SRF)-mediated transcription contributes to developmental and adult brain plasticity. Therefore, we investigated the role of a newly identified SRF coactivator, MKL1, in the regulation of SRF-driven transcription in rat forebrain neurons. MKL1 expression was found in newborn rat cortical or hippocampal neurons in culture as well as in adult rat forebrain. Immunostaining demonstrated constitutive nuclear localization of MKL1 in the CA1 region of the hippocampus, in the deep layers of the neocortex, and in cultured neurons. Overexpression of MKL1 in primary cortical neurons elevated SRF-driven transcription and enhanced its stimulation by BDNF. In addition, inhibition of endogenous MKL1 by overexpression of a dominant-negative MKL1 mutant or by small interfering RNA reduced BDNF activation of SRF-driven transcription. In neurons, endogenous MKL1 was associated with SRF-regulated chromatin regions of several endogenous genes including c-fos, JunB, Srf, and Cyr61. BDNF activation of MKL1/SRF-driven transcription was dependent on the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway, which also led to MKL1 phosphorylation. Finally, synaptic activity stimulation of SRF-driven transcription was reduced by inhibition of endogenous MKL1. Conversely, synaptic activity enhanced transcription by overexpressed MKL1. MKL1 regulation by synaptic activity was mediated through the NMDA receptor-activated ERK1/2. These results suggest that neuronal MKL1 contributes to SRF-regulated gene expression induced by BDNF or synaptic activity. In addition, MKL1 appears as a novel mediator of the signaling between ERK1/2 and SRF. Moreover, MKL1 is a likely regulator of SRF-driven transcription programs that underlie neuronal plasticity.

Although the transcription factor serum response factor (SRF) has been suggested to play a role in activity-dependent gene expression and mediate plasticity-associated structural changes in the hippocampus, no unequivocal evidence has been provided for its role in brain pathology, such as epilepsy. A genome-wide program of activity-induced genes that are regulated by SRF also remains unknown. In the present study, we show that the inducible and conditional deletion of SRF in the adult mouse hippocampus increases the epileptic phenotype in the kainic acid model of epilepsy, reflected by more severe and frequent seizures. Moreover, we observe a robust decrease in activity-induced gene transcription in SRF knockout mice. We characterize the genetic program controlled by SRF in neurons and using functional annotation, we find that SRF target genes are associated with synaptic plasticity and epilepsy. Several of these SRF targets function as regulators of inhibitory or excitatory balance and the structural plasticity of neurons. Interestingly, mutations in those SRF targets have found to be associated with such human neuropsychiatric disorders, as autism and intellectual disability. We also identify novel direct SRF targets in hippocampus: Npas4, Gadd45g, and Zfp36. Altogether, our data indicate that proteins that are highly upregulated by neuronal stimulation, identified in the present study as SRF targets, may function as endogenous protectors against overactivation. Thus, the lack of these effector proteins in SRF knockout animals may lead to uncontrolled excitation and eventually epilepsy.Electronic supplementary materialThe online version of this article (doi:10.1007/s12035-014-9089-7) contains supplementary material, which is available to authorized users.