A subset of genes implicated in genetic and acquired neurological disorders encode proteins essential to neural patterning and neurogenesis. The gene silencing transcription factor neuronal repressor element-1 silencing transcription factor (REST)/neuron-restrictive silencer factor (NRSF) plays a critical role in elaboration of the neuronal phenotype. In neural progenitor and non-neural cells, REST acts by repression of a subset of neural genes important to synaptic plasticity and synaptic remodeling, including the AMPA receptor (AMPAR) subunit GluR2. Here we show that global ischemia triggers REST mRNA and protein expression. REST suppresses GluR2 promoter activity and gene expression in neurons destined to die. Because the GluR2 subunit governs AMPAR Ca2+ permeability, these changes are expected to have profound effects on neuronal survival. In keeping with this concept, acute knockdown of the REST gene by antisense administration prevents GluR2 suppression and rescues post-ischemic neurons from ischemia-induced cell death in an in vitro model. To our knowledge, our study represents the first example of ischemia-induced induction of a master transcriptional regulator gene and its protein expression critical to neural differentiation and patterning in adult neurons. Derepression of REST is likely to be an important mechanism of insult-induced neuronal death.

Transient global ischemia is a neuronal insult that induces delayed

Mature oocyte cytoplasm can reprogram somatic cell nuclei to the pluripotent state through a series of sequential events including protein exchange between the donor nucleus and ooplasm, chromatin remodeling, and pluripotency gene reactivation. Maternal factors that are responsible for this reprogramming process remain largely unidentified. Here, we demonstrate that knockdown of histone variant H3.3 in mouse oocytes results in compromised reprogramming and down-regulation of key pluripotency genes; and this compromised reprogramming for developmental potentials and transcription of pluripotency genes can be rescued by injecting exogenous H3.3 mRNA, but not H3.2 mRNA, into oocytes in somatic cell nuclear transfer embryos. We show that maternal H3.3, and not H3.3 in the donor nucleus, is essential for successful reprogramming of somatic cell nucleus into the pluripotent state. Furthermore, H3.3 is involved in this reprogramming process by remodeling the donor nuclear chromatin through replacement of donor nucleus-derived H3 with de novo synthesized maternal H3.3 protein. Our study shows that H3.3 is a crucial maternal factor for oocyte reprogramming and provides a practical model to directly dissect the oocyte for its reprogramming capacity. P ioneering nuclear transfer experiments in amphibians have revealed that the cytoplasm of the egg is able to reprogram a differentiated nucleus to the embryonic state (1, 2). The success of somatic cell nuclear transfer (SCNT) to produce cloned animals using enucleated metaphase II (MII) oocytes (3, 4), and, recently, the successful derivation of SCNT human embryonic stem cells (5), have demonstrated that maternal factors in the mature ooplasm are capable and sufficient to reprogram a differentiated cell nucleus to pluripotency. This process is known to involve a series of sequential events including protein exchange between donor nucleus and ooplasm, donor nuclear chromatin remodeling, and pluripotency gene reactivation (6-12). However, maternal factors responsible for this reprogramming process and the underlying mechanism(s) remain largely unknown.Thousands of different maternal proteins and mRNAs have been found in mouse mature oocytes (13,14), including variants of the core histone proteins that, along with DNA, constitute nucleosomes. Accumulating evidence suggests that histone variants play important roles in chromatin remodeling and epigenetic regulation orchestrating gene expression changes during reprogramming (12,15,16). In mammals, the histone variant H3.3 is encoded by two different genes (h3f3a and h3f3b), whose translation results in an identical protein product (17, 18). Unlike canonical H3 histones that are expressed and incorporated into chromatin during S phase, expression of H3.3 is not cell cycle-regulated, and the variant is expressed in quiescent cells, postmitotic cells, and proliferating cells throughout the whole cycle, enabling H3.3 deposition in a DNA synthesis-independent manner during and outside of S phase (19). It has been suggested that matern...

Transient forebrain or global ischemia induces delayed neuronal death in vulnerable CA1 pyramidal cells with many features of apoptosis. A brief period of ischemia, i.e., ischemic preconditioning, affords robust protection of CA1 neurons against a subsequent more prolonged ischemic challenge. Here we show that preconditioning acts via PI3K/Akt signaling to block the ischemia-induced cascade involving mitochondrial translocation of Bad, assembly of Bad with Bcl-x L , cleavage of Bcl-x L to form its prodeath fragment, ΔN-Bcl-x L , activation of large-conductance channels in the mitochondrial outer membrane, mitochondrial release of cytochrome c and Smac/DIABLO (second mitochondria-derived activator of caspases/direct IAP-binding protein with low pI), caspase activation, and neuronal death. These findings show how preconditioning acts to prevent the release of cytochrome c and Smac/DIABLO from mitochondria and to preserve the integrity of the mitochondrial membrane. The specific PI3K inhibitor LY294002 administered in vivo 1 h before or immediately after ischemia or up to 120 h later significantly reverses preconditioning-induced protection, indicating a requirement for sustained PI3K signaling in ischemic tolerance. These findings implicate PI3K/Akt signaling in maintenance of the integrity of the mitochondrial outer membrane.