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S-Nitrosoglutathione reductase (GSNOR) is a key regulator of protein S-nitrosylation, the covalent modification of cysteine residues by nitric oxide that can affect activities of many proteins. We recently discovered that excessive S-nitrosylation from GSNOR deficiency in mice under inflammation inactivates the key DNA repair protein O(6) -alkylguanine-DNA alkyltransferase and promotes both spontaneous and carcinogen-induced hepatocellular carcinoma. To explore further the mechanism of tumorigenesis due to GSNOR deficiency, we compared the protein expression profiles in the livers of wild-type and GSNOR-deficient (GSNOR(-/-) ) mice that were challenged with lipopolysaccharide to induce inflammation and expression of inducible nitric oxide synthase (iNOS). Two-dimensional difference gel electrophoresis analysis identified 38 protein spots of significantly increased intensity and 31 protein spots of significantly decreased intensity in the GSNOR(-/-) mice compared to those in the wild-type mice. We subsequently identified 19 upregulated and 19 downregulated proteins in GSNOR(-/-) mice using mass spectrometry. Immunoblot analysis confirmed in GSNOR(-/-) mice a large increase in the expression of the pro-inflammatory mediator S100A9, a protein previously implicated in human liver carcinogenesis. We also found a decrease in the expression of multiple members of the protein disulfide-isomerase (PDI) family and an alteration in the expression pattern of the endoplasmic reticulum (ER) chaperones in GSNOR(-/-) mice. Furthermore, altered expression of these proteins from GSNOR deficiency was prevented in mice lacking both GSNOR and iNOS. In addition, we detected S-nitrosylation of two members of the PDI protein family. These results suggest that S-nitrosylation resulting from GSNOR deficiency may promote carcinogenesis under inflammatory conditions in part through the disruption of inflammatory and ER stress responses.

Abstract.: MicroRNAs (miRNAs) are endogenous small noncoding RNAs (20 – 23 nucleotides) that negatively regulate the gene expressions at the posttranscriptional level by base pairing to the 3′ untranslated region of target messenger RNAs. Hundreds of miRNAs have been identified in humans and evolutionarily conserved from plants to animals. It is revealed that miRNAs regulate various physiological and pathological pathways such as cell differentiation, cell proliferation, and tumoriogenesis. By the computational analysis, it is predicted that 30% of protein-encoding genes are regulated by miRNAs. In this review, we discuss recent remarkable advances in the miRNA biogenetic and functional mechanisms and the involvements of miRNAs in cell differentiation, especially in hematopoietic lineages, and cancer. These evidences offer the possibility that miRNAs would be potentially useful for drug discovery. Keywords:: microRNA, RNA cleavage, translational repression, target mRNA, base pairing

Abstract.: P2X receptors belong to a unique family of ligand-gated channels in terms of their molecular architecture, in which the channel subunit has two transmembrane alpha-helixes with a large extracellular loop keeping amino- and carboxy-termini in the cytoplasm. Post-transcriptional modifications of P2X receptors could diversify cellular responsiveness induced by extracellular ATP in anterior pituitary cells and other cell types. Recently, we found a spliced variant P2X2 transcript, termed P2X2e, in mouse pituitary. The P2X2e has a shorter cytoplasmic carboxy-terminal tail than those of full-length P2X2a or splice variant P2X2b subunits. Although ATP induced rapid responses in all homomeric P2X2 channels, the current induced by P2X2e declined significantly faster than those by P2X2a or P2X2b. In this article, we summarize functional alterations of P2X2 receptors after splicing reactions. Combinations of different P2X2 subunit carboxy-termini to form homomeric and heteromeric channels could be a molecular mechanism for promoting functional diversities of ATP-induced cellular signals. Keywords:: purinergic receptor, P2X2, splicing, protein interaction, mutagenesis

Hypertension is a complex trait, influenced by multiple genetic and environmental factors (1). Although it has been estimated that approximately 30-50% of blood pressure variance is due to inherited genes, [...], Malfunction of the circadian clock has been linked to the pathogenesis of a variety of diseases. We show that mice lacking the core clock components Cryptochrome-1 (Cry1) and Cryptochrome-2 (Cry2) (Cry-null mice) show salt-sensitive hypertension due to abnormally high synthesis of the mineralocorticoid aldosterone by the adrenal gland. An extensive search for the underlying cause led us to identify type VI 3β-hydroxyl-steroid dehydrogenase (Hsd3b6) as a new hypertension risk factor in mice. Hsd3b6 is expressed exclusively in aldosterone-producing cells and is under transcriptional control of the circadian clock. In Cry-null mice, Hsd3b6 messenger RNA and protein levels are constitutively high, leading to a marked increase in 3(3- hydroxysteroid dehydrogenase-isomerase (3β-HSD) enzymatic activity and, as a consequence, enhanced aldosterone production. These data place Hsd3b6 in a pivotal position through which circadian clock malfunction is coupled to the development of hypertension. Translation of these findings to humans will require clinical examination of human HSD3B1 gene, which we found to be functionally similar to mouse Hsd3b6.

[[Arg.sup.8]]-vasopressin (Avp), a nonapeptide hormone, is known to regulate blood pressure, water balance, and a variety of behaviors such as anxiety, aggression, and bonding. Although some evidence that Avp modifies ethanol consumption and some of the effects of ethanol on behavior have been reported, the role of Avp in alcohol consumption and preference is poorly understood. The Avp1a receptor (Avpr1a) is ubiquitously expressed in the central nervous system. To determine the role of Avp signaling on the behavioral effects of alcohol, we examined voluntary ethanol consumption in mice with targeted disruptions of the Avpr1a knockout (Avpr1a KO) gene. Avpr1a KO mice displayed both increased ethanol consumption and preference compared with wild-type (WT) mice. Enhanced ethanol consumption was dramatically and reversibly reduced by treatment with N-methyl-D-aspartic acid antagonists. Basal glutamate release was elevated around the striatum in Avpr1a KO mice. Elevation of extracellular glutamate was also produced in WT mice by local application of an Avpr1a antagonist though a dialysis probe, and this elevation was quickly reversed by stopping the perfusion. These results suggest that Avp can inhibit the release of glutamate from the presynaptic terminal via the Avp1a receptor and that elevation of glutamate levels owing to loss of the inhibitory effect via Avp-Avpr1a signaling may play an important role in the preference for ethanol. [[Arg.sup.8]]-vasopressin

Attenuation of early restenosis after percutaneous coronary intervention (PCI) is important for the successful treatment of coronary artery disease. Some clinical studies have shown that hypertension is a risk factor for early restenosis after PCI. These findings suggest that [[alpha].sub.1]-adrenergic receptors ([[alpha].sub.1]-ARs) may facilitate restenosis after PCI because of [[alpha].sub.1]-AR's remarkable contribution to the onset of hypertension. In this study, we examined the neointimal formation after vascular injury in the femoral artery of [[alpha].sub.1A]-knockout ([[alpha].sub.1A]-KO), [[alpha].sub.1B]-KO, [[alpha].sub.1D]-KO, [[alpha].sub.1A]-/[[alpha].sub.1B]-AR double-KO ([[alpha].sub.1AB]-KO), and wild-type mice to investigate the functional role of each [[alpha].sub.1]-AR subtype in neointimal formation, which is known to promote restenosis. Neointimal formation 4 wk after wire injury was significantly (P < 0.05) smaller in [[alpha].sub.1AB]-KO mice than in any other group of mice, while blood pressures were not altered in any of the groups of mice after wire injury compared with those before it. These results suggest that lack of both [[alpha].sub.1A]- and [[alpha].sub.1B]-ARs could be necessary to inhibit neointimal formation in the mouse femoral artery. [[alpha].sub.1]-adrenergic receptor: vascular injury; smooth muscle cell: femoral artery of mouse; wire injury doi:10.1152/ajpheart.00626.2006

Excessive proliferation of vascular wall cells underlies the development of elevated vascular resistance in hypoxic pulmonary hypertension (PH), but the responsible mechanisms remain unclear. Growth-promoting effects of catecholamines may contribute. Hypoxemia causes sympathoexcitation, and prolonged stimulation of [[alpha].sub.1]-adrenoceptors ([[alpha].sub.1]-ARs) induces hypertrophy and hyperplasia of arterial smooth muscle cells and adventitial fibroblasts. Catecholamine trophic actions in arteries are enhanced when other conditions favoring growth or remodeling are present, e.g., injury or altered shear stress, in isolated t pulmonary arteries from rats with hypoxic PH. The present study examined the hypothesis that catecholamines contribute to pulmonary vascular remodeling in vivo in hypoxic PH. Mice genetically deficient in norepinephrine and epinephrine production [dopamine [beta]-[hydroxylase.sup.-/-] (DB[H.sup.-/-])] or [[alpha].sub.1]-ARs were examined for alterations in PH, cardiac hypertrophy, and vascular remodeling after 21 days exposure to normobaric 0.1 inspired oxygen fraction ([MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII.]). A decrease in the lumen area and an increase in the wall thickness of arteries were strongly inhibited in knockout mice (order of extent of inhibition: DB[H.sup.-/-] = [[alpha].sub.1D]-A[R.sup.-/-] > [[alpha].sub.1B]-A[R.sup.-/-]). Distal muscularization of small arterioles was also reduced (DB[H.sup.-/-] > [[alpha].sub.1D]-A[R.sup.-/-] > [[alpha].sub.1B]-A[R.sup.-/-] mice). Despite these reductions, increases in right ventricular pressure and hypertrophy were not attenuated in DB[H.sup.-/-] and [[alpha].sub.1B]-A[R.sup.-/-] mice. However, hematocrit increased more in these mice, possibly as a consequence of impaired cardiovascular activation that t occurs during reduction of [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII.]. In contrast, in [[alpha].sub.1D]-A[R.sup.-/-] mice, where hematocrit increased the same as in wild-type mice, right ventricular pressure was reduced. These data suggest that catecholamine stimulation of [[alpha].sub.1B]- and [[alpha].sub.1D]-ARs contributes significantly to vascular remodeling in hypoxic PH. vascular smooth muscle; adrenergic receptors; hypoxia doi:10.1152/ajpheart.00792.2006