Enhancement of cerebral blood flow by hypoxia is critical for brain function, but signaling systems underlying its regulation have been unclear. We report a pathway mediating hypoxia-induced cerebral vasodilation in studies monitoring vascular disposition in cerebellar slices and in intact mouse brains using two-photon intravital laser scanning microscopy. In this cascade, hypoxia elicits cerebral vasodilation via the coordinate actions of H 2 S formed by cystathionine β-synthase (CBS) and CO generated by heme oxygenase (HO)-2. Hypoxia diminishes CO generation by HO-2, an oxygen sensor. The constitutive CO physiologically inhibits CBS, and hypoxia leads to increased levels of H 2 S that mediate the vasodilation of precapillary arterioles. Mice with targeted deletion of HO-2 or CBS display impaired vascular responses to hypoxia. Thus, in intact adult brain cerebral cortex of HO-2-null mice, imaging mass spectrometry reveals an impaired ability to maintain ATP levels on hypoxia.gas biology | neurovascular unit | energy metabolism | gasotransmitter T he cerebral circulation is maintained by autoregulation, which prevents marked alterations in response to changes in blood pressure, whereas functional hyperemia links blood flow to neural activity (1). Blood flow regulation in the brain is modulated by O 2 (2), with increased cerebral blood flow in response to hypoxia critical for protecting the brain against diverse insults. Such regulation also participates in functional hyperemia, as demonstrated by functional MRI investigations indicating a transient decrease in O 2 levels preceding activation of blood flow in response to neuronal firing (3).Alterations in cerebral blood flow in response to hypoxia and neural activity are mediated via several neurotransmitter systems, with prominent involvement of the gaseous mediator nitric oxide (NO) (1, 2). In response to glutamate acting on NMDA receptors, neuronal NO synthase (nNOS) is activated by increases in intracellular calcium, with the generated NO stimulating soluble guanylyl cyclase, thereby increasing cGMP levels to dilate blood vessels (4). Functional hyperemia is decreased by ∼50% in rats in response to inhibition of nNOS (5). Another gaseous mediator, CO (6-8), is also vasoactive. In some blood vessel systems (e.g., liver sinusoids), CO causes vasodilation, and inhibition of its biosynthetic enzyme HO-2 leads to vasoconstriction (9-13). However, in the cerebral circulation, CO elicits vasoconstriction. Thus, HO inhibitors cause cerebral vasodilation, an effect reversed by CO (14). This action of CO cannot be readily explained by previously identified CO receptors, such as soluble guanylyl cyclase (6-12, 15) or potassium channels (13, 16), both of which mediate vasodilation. The CO and NO systems interface; thus, the vasodilatory actions of HO inhibitors are partially reversed by inhibitors of NOS (14). A third gaseous mediator, H 2 S, is also vasoactive, eliciting vasodilation in both the peripheral and cerebral circulation (17-21). H 2 S can be physiologically ...

The core histones are essential components of the nucleosome that act as global negative regulators of DNA-mediated reactions including transcription, DNA replication and DNA repair. Modified residues in the N-terminal tails are well characterized in transcription, but not in DNA replication and DNA repair. In addition, roles of residues in the core globular domains are not yet well characterized in any DNA-mediated reactions. To comprehensively understand the functional surface(s) of a core histone, we constructed 320 yeast mutant strains, each of which has a point mutation in a core histone, and identified 42 residues responsible for the suppressor of Ty (Spt -) phenotypes, and 8, 30 and 61 residues for sensitivities to 6-azauracil (6AU), hydroxyurea (HU) and methyl-methanesulfonate (MMS), respectively. In addition to residues that affect one specific assay, residues involved in multiple reactions were found, and surprisingly, about half of them were clustered at either the nucleosome entry site, the surface required for nucleosome-nucleosome interactions in crystal packing or their surroundings. This comprehensive mutation approach was proved to be powerful for identification of the functional surfaces of a core histone in a variety of DNA-mediated reactions and could be an effective strategy for characterizing other evolutionarily conserved hub-like factors for which surface structural information is available.

Post-translational modification of histones, a major protein component of eukaryotic chromosomes, contributes to the epigenetic regulation of gene expression. Distinct patterns of histone modification are observed at specific chromosomal regions and affect various reactions on chromosomes (transcription, replication, repair, and recombination). Histone modification has long been proposed to have a profound effect on eukaryotic gene expression since its discovery in 1964. Verification of this idea, however, was difficult until the identification of enzymes responsible for histone modifications. Ten years ago (1995), histone acetyltransferases (HATs), which acetylate lysine residues in histone amino-terminal tail regions, were isolated. HATs are involved in the regulation of both promoter-specific transcription and long-range/chromosome-wide transcription. Analyses of HATs and other modification enzymes have revealed mechanisms of epigenetic regulation that are mediated by post-translational modifications of histones. Here we review some major advances in the field, with emphasis on the lysine specificity of the acetylation reaction and on the regulation of gene expression over broad regions.

The attachment of sister kinetochores to microtubules from opposite spindle poles is essential for faithful chromosome segregation. Kinetochore assembly requires centromere‐specific nucleosomes containing the histone H3 variant CenH3. However, the functional roles of the canonical histones (H2A, H2B, H3, and H4) in chromosome segregation remain elusive. Using a library of histone point mutants in Saccharomyces cerevisiae, 24 histone residues that conferred sensitivity to the microtubule‐depolymerizing drugs thiabendazole (TBZ) and benomyl were identified. Twenty‐three of these mutations were clustered at three spatially separated nucleosomal regions designated TBS‐I, ‐II, and ‐III (TBZ/benomyl‐sensitive regions I–III). Elevation of mono‐polar attachment induced by prior nocodazole treatment was observed in H2A‐I112A (TBS‐I), H2A‐E57A (TBS‐II), and H4‐L97A (TBS‐III) cells. Severe impairment of the centromere localization of Sgo1, a key modulator of chromosome bi‐orientation, occurred in H2A‐I112A and H2A‐E57A cells. In addition, the pericentromeric localization of Htz1, the histone H2A variant, was impaired in H4‐L97A cells. These results suggest that the spatially separated nucleosomal regions, TBS‐I and ‐II, are necessary for Sgo1‐mediated chromosome bi‐orientation and that TBS‐III is required for Htz1 function.