Autophagy is a process by which cytoplasmic organelles can be catabolized either to remove defective structures or as a means of providing macromolecules for energy generation under conditions of nutrient starvation. In this study we demonstrate that mitochondrial autophagy is induced by hypoxia, that this process requires the hypoxia-dependent factor-1-dependent expression of BNIP3 and the constitutive expression of Beclin-1 and Atg5, and that in cells subjected to prolonged hypoxia, mitochondrial autophagy is an adaptive metabolic response which is necessary to prevent increased levels of reactive oxygen species and cell death.The survival of metazoan organisms is dependent upon their ability to efficiently generate energy through the process of mitochondrial oxidative phosphorylation in which reducing equivalents, derived from the oxidation of acetyl CoA in the tricarboxylic acid cycle, are transferred from NADH and FADH 2 to the electron transport chain and ultimately to O 2 , a process which produces an electrochemical gradient that is used to synthesize ATP (1). Although oxidative phosphorylation is more efficient than glycolysis in generating ATP, it carries the inherent risk of generating reactive oxygen species (ROS) 2 as a result of electrons prematurely reacting with O 2 at respiratory complex I or complex III. Transient, low level ROS production is utilized for signal transduction in metazoan cells, but prolonged elevations of ROS result in the oxidation of protein, lipid, and nucleic acid leading to cell dysfunction or death.O 2 delivery and utilization must, therefore, be precisely regulated to maintain energy and redox homeostasis.Hypoxia-inducible factor 1 (HIF-1) plays a key role in the regulation of oxygen homeostasis (2, 3). HIF-1 is a heterodimer composed of a constitutively expressed HIF-1␤ subunit and an O 2 -regulated HIF-1␣ subunit (4). Under aerobic conditions, HIF-1␣ is hydroxylated on proline residue 402 and/or 564 by prolyl hydroxylase 2 a dioxygenase that utilizes O 2 and ␣-ketoglutarate as co-substrates with ascorbate as co-factor in a reaction that generates succinate and CO 2 as side products (5-8). Under hypoxic conditions the rate of hydroxylation declines, either as a result of inadequate substrate (O 2 ) or as a result of hypoxia-induced mitochondrial ROS production, which may oxidize Fe(II) in the catalytic center of the hydroxylase (9, 10). Hydroxylated HIF-1␣ is bound by the von HippelLindau protein, which recruits a ubiquitin protein ligase complex that targets HIF-1␣ for proteasomal degradation (11)(12)(13)(14).HIF-1 regulates the transcription of hundreds of genes in response to hypoxia (15, 16), including the EPO (17) and VEGF (18) genes that encode proteins required for erythropoiesis and angiogenesis, respectively, which serve to increase O 2 delivery. In addition, HIF-1 controls a series of molecular mechanisms designed to maintain energy and redox homeostasis. First, HIF-1 coordinates a switch in the composition of cytochrome c oxidase (mitochondrial electron-transp...

A library of drugs that are in clinical trials or use was screened for inhibitors of hypoxia-inducible factor 1 (HIF-1). Twenty drugs inhibited HIF-1-dependent gene transcription by >88% at a concentration of 0.4 M. Eleven of these drugs were cardiac glycosides, including digoxin, ouabain, and proscillaridin A, which inhibited HIF-1␣ protein synthesis and expression of HIF-1 target genes in cancer cells. Digoxin administration increased latency and decreased growth of tumor xenografts, whereas treatment of established tumors resulted in growth arrest within one week. Enforced expression of HIF-1␣ by transfection was not inhibited by digoxin, and xenografts derived from these cells were resistant to the anti-tumor effects of digoxin, demonstrating that HIF-1 is a critical target of digoxin for cancer therapy.cancer therapy ͉ hypoxia ͉ tumor xenograft

We report a conceptually new approach to the direct amination of aromatic C-H bonds. In this process, an oxime ester function reacts with an aromatic C-H bond under redox-neutral conditions to form, in the case studied, an indole product. These reactions occur with relatively low catalyst loading (1 mol %) by a mechanism that appears to involve an unusual initial oxidative addition of an N-O bond to a Pd(0) species. The Pd(II) complex from oxidative addition of the N-X bond has been isolated for the first time, and evidence for the intermediacy of such oxidative addition products in the catalytic reaction has been gained.

Palladium-catalyzed direct arylations of benzene have been proposed to occur by the generation of a phosphine-ligated arylpalladium pivalate complex LPd(Ar)(OPiv) and reaction of this complex with benzene. We have isolated an example of the proposed intermediate and evaluated whether this complex reacts with benzene to form the biaryl products of the catalytic process. In contrast to the proposed mechanism, no biaryl product was formed from cleavage of the benzene C-H bond by LPd(Ar)(OPiv). However, reactions of LPd(Ar)OPiv with benzene and additives that displace or consume the phosphine ligand formed the arylated products in good yield, suggesting that a "ligandless" arylpalladium(II) carboxylate complexes undergoes the C-H cleavage step. Consistent with this conclusion, we found that reactions catalyzed by Pd(OAc) 2 without ligand occur faster than, and with comparable selectivities to, reactions catalyzed by Pd(OAc) 2 and phosphine ligand.Direct arylation, the reaction of aryl halides with arenes or heteroarenes to form biaryl or aryl-heteroaryl products, is an attractive alternative to traditional cross-coupling because it occurs without the need to prepare organometallic or main-group reagents. 1 Early efforts to develop this process focused on the direct arylation of heteroarenes, 2 arenes with directing groups 3 and electron-deficient arenes. 4 In 2006, the direct arylation of benzene was reported by Lafrance and Fagnou. 5 Substoichiometric amounts of pivalic acid were used in the reported catalyst system, and this carboxylic acid was proposed to function as a proton shuttle during the aryl C-H cleavage step. 6 Despite the improvements in the reaction scope, little experimental information on the mechanism of this reaction is available, especially on the mechanism of the cleavage of the C-H bonds in arenes. Phosphine-ligated arylpalladium carboxylates LPd(Ar)(OCOR) are typically proposed to react with heteroarenes or arenes to form biarylpalladium complexes through a concerted metallation-deprotonation (CMD) pathway. 7 DFT calculations of this pathway have been conducted, and these studies suggested that arylpalladium acetates are competent to undergo C-H cleavage with various heteroarenes.8 Indeed, a recent report from Fagnou's group showed that the reaction of Pd(PtBu 3 )(Ph)(OAc) with 4-nitropyridine Noxide formed the arylated product. 9 However, arylpalladium pivalate complexes have not been studied either experimentally or theoretically, and the reactions of isolated arylpalladium carboxylate complexes with arenes have not been conducted. We report the synthesis and characterization of the phosphine-ligated arylpalladium pivalate complexes Pd(PtBu 3 )(Ar)(OPiv). Inconsistent with previous proposals, the reaction of isolated Pd(PtBu 3 )(Ar)(OPiv) with benzene does not form Ar-Ph products. However, reactions conducted with additives that displace or consume the phosphine ligand promote the formation of the arylated products, and these results and others reported herein imply the involvement o...