Manipulation of redox signaling in mammalian cells enabled by controlled photogeneration of reactive oxygen species

Posen, Yehudit; Kalchenko, Vyacheslav; Seger, Rony; Brandis, Alexander; Scherz, Avigdor; Salomon, Yoram

doi:10.1242/jcs.02323

Cited by 39 publications

(36 citation statements)

References 64 publications

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“…Previously, this was attributed to the reaction between · OH and HE [32]. However, we found that SOD vastly diminished 2-OH-E + formation, suggesting that superoxide is responsible for 2-OH-E + generation in this system.…”

Section: Discussionmentioning

confidence: 49%

Pulse radiolysis and steady-state analyses of the reaction between hydroethidine and superoxide and other oxidants

Zielonka¹,

Sarna²,

Roberts³

et al. 2006

Archives of Biochemistry and Biophysics

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Hydroethidine (HE) is a cell-permeable probe used for the intracellular detection of superoxide. Here we report the direct measurement of the rate constant between hydroethidine and superoxide radical anion using the pulse radiolysis technique. This reaction rate constant was calculated to be ca. 2·10 6 M -1 s -1 in water:ethanol (1:1) mixture. The spectral characteristics of the intermediates indicated that the one-electron oxidation product of HE was different from the one-electron reduction product of ethidium (E + ). The HPLC-electrochemical measurements of incubation mixtures containing HE and the oxygenated Fenton's reagent (Fe 2+ /DTPA/H 2 O 2 ) in the presence of aliphatic alcohols or formate as a superoxide generating system revealed 2-OH-E + as a major product. Formation of 2-OH-E + by the Fenton's reagent without additives was shown to be superoxide dismutase-sensitive and we attribute the formation of superoxide radical anion to the one-electron reduction of oxygen by the DTPA-derived radical. Addition of tert-butanol, DMSO, and potassium bromide to the Fenton's system caused inhibition of 2-OH-E + formation. Results indicate that reducing and oxidizing radicals have differential effects on the formation of 2-OH-E + .

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Section: Discussionmentioning

confidence: 49%

Pulse radiolysis and steady-state analyses of the reaction between hydroethidine and superoxide and other oxidants

Zielonka¹,

Sarna²,

Roberts³

et al. 2006

Archives of Biochemistry and Biophysics

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“…Indeed, such a process has been demonstrated for both JNK1/2 (18,19) and p38␣/␤ (20,21) in some systems. However, in other studies, it was suggested that JNK1/2 (22, 23) and p38␣/␤ (24,25) are in fact localized in the nucleus of resting cells. For the latter, it was even shown that after stimulation it is exported out of the nucleus (26).…”

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confidence: 90%

Beta-Like Importins Mediate the Nuclear Translocation of Mitogen-Activated Protein Kinases

Zehorai

Seger

2014

Molecular and Cellular Biology

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The rapid nuclear translocation of signaling proteins upon stimulation is important for the regulation of de novo gene expression. We have studied the stimulated nuclear shuttling of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinases (MAPKs) and found that they translocate into the nucleus in a Ran-dependent, but NLS-or NTS-independent, manner, unrelated to their catalytic activity. We show that this translocation involves three ␤-like importins, importins 3, 7, and 9 (Imp3/ 7/9). Knockdown of these importins inhibits the nuclear translocation of the MAPKs and, thereby, activation of their transcription factor targets. We further demonstrate that the translocation requires the stimulated formation of heterotrimers composed of Imp3/Imp7/MAPK or Imp3/Imp9/MAPK. JNK1/2 and p38␣/␤ bind to either Imp7 or Imp9 upon stimulated posttranslational modification of the two Imps, while Imp3 joins the complex after its stimulation-induced phosphorylation. Once formed, these heterotrimers move to the nuclear envelope, where importin 3 remains, while importins 7 and 9 escort the MAPKs into the nucleus. These results suggest that ␤-like importins are central mediators of stimulated nuclear translocation of signaling proteins and therefore add a central level of regulation to stimulated transcription. Rapid and massive nuclear translocation of signaling proteins is an important step in the induction of transcription upon extracellular stimulation (1). Despite the importance of this process, its molecular mechanism has only been elucidated for a few signaling components. The well-understood cases include signaling proteins such as NF-B (2) and extracellular signal-regulated kinase 5 (ERK5) (3), which utilize the classical nuclear localization signal (NLS)-mediated binding with importin-␣ (Imp␣) and Imp␤ (4). However, many other signaling proteins (Ͼ60) translocate to the nucleus shortly after stimulation by using distinct NLS-and Imp␣/␤-independent mechanisms. The rapid nature of the nuclear translocation and size of the shuttling proteins make it unlikely that their translocation is mediated by passive diffusion. Several other stimulus-dependent signaling mechanisms for such proteins that have been proposed include Imp-like activity by the shuttled protein itself (e.g., ␤-catenin [5]) or the use of shuttling proteins other than Imp␣/␤ (e.g., Smad4 [6]). We recently elucidated the molecular mechanism of ERK1/2 translocation, showing that it involves casein kinase 2 (CK2)-mediated phosphorylation of two Ser residues within a nuclear translocation signal (NTS) of ERK1/2 (7,8). This phosphorylation allows interaction with Imp7, which further facilitates ERK1/2 shuttling via nuclear pores. Aside from ERK1/2, NTS-like sequences have been seen in only a few other NLS-deficient translocating proteins (SMAD3, MEK1 [7], and Egr1 [9]), indicating that other proteins use a distinct mechanism for their translocation.The involvement of Imp7 in the nuclear translocation of ERK1/2 drew our attention to the group of ill-studi...

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“…While the mechanism for TH preservation of Akt phosphorylation is unknown, it could potentially alter the relative balance of kinase or phosphatase activity. Alternatively, TH could modulate Akt activity through the activation of heat shock/cold shock proteins or the modulation of reactive oxygen species-sensitive signaling pathways (10,35,39). The observed reduction in Akt phosphorylation in Akt1 ϩ/Ϫ mice could reflect a change in the relative stoichiometry of Akt, kinases, and phosphatases including protein phosphatase 2 in these animals.…”

Section: Discussionmentioning

confidence: 99%

Akt1 genetic deficiency limits hypothermia cardioprotection following murine cardiac arrest

Beiser

Wojcik

Zhao

et al. 2010

American Journal of Physiology-Heart and Circulatory Physiology

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Beiser DG, Wojcik KR, Zhao D, Orbelyan GA, Hamann KJ, Vander Hoek TL. Akt1 genetic deficiency limits hypothermia cardioprotection following murine cardiac arrest. Therapeutic hypothermia (TH) cardioprotection has recently been associated with increased Akt signaling in a rat model of cardiac arrest. However, it is not known whether Akt is required for this beneficial effect of TH. We used a mouse model of cardiac arrest demonstrating TH cardioprotection to study the response of mice deficient in an Akt1 allele. We hypothesized that Akt1 mediates TH cardioprotection and that decreases in Akt1 content would diminish such protection. Adult C57BL/6 wildtype (WT) mice underwent an 8-min cardiac arrest. After 6 min, the mice were randomized to normothermia (WTNT, 37°C) or TH (WTTH, 30°C). Following cardiopulmonary resuscitation and the return of spontaneous circulation (ROSC), the animals were hemodynamically monitored for 240 min (R240). At R240, cardiac tissue Akt content and phosphorylation were assayed. Studies were repeated in Akt1 heterozygous (Akt1 ϩ/Ϫ ) mice. As a result, baseline characteristics and ROSC rates were equivalent across groups. At R240, WTTH mice exhibited lower heart rate, larger stroke volume, and higher cardiac output than WTNT animals (P Ͻ 0.05). Cardioprotection in WTTH at R240 was associated with increased cardiac Akt phosphorylation at Ser473 and Thr308 compared with that in WTNT (P Ͻ 0.05). THassociated alterations in Akt phosphorylation, stroke volume, heart rate, and cardiac output were abrogated in Akt1 ϩ/Ϫ animals. In conclusion, TH improves post-ROSC cardiac function and increases Akt phosphorylation in WT, but not Akt1 ϩ/Ϫ , mice. The Akt1 isoform appears necessary for TH-mediated cardioprotection.sudden cardiac death; asystole; cardiopulmonary resuscitation; therapeutic hypothermia; protein kinase B SUDDEN CARDIAC ARREST is a leading contributor to cardiovascular disease mortality with only 5-7% of out-of-hospital cardiac arrest victims surviving hospital discharge (27). During the first minutes to hours following the return of spontaneous circulation (ROSC), patients often exhibit severe and lethal myocardial dysfunction than can occur in the absence of underlying focal coronary artery occlusion (8,16,25,38,41). The mechanisms of post-ROSC myocardial depression are likely related to factors associated with ischemia-reperfusion (I/R) injury including substrate depletion, acidosis, oxidative stress, mitochondrial injury, alterations in intracellular calcium handling and in circulating cytokines, and the posttranslational modification of contractile and mitochondrial proteins (6,20,23,24,28).The induction of mild to moderate therapeutic hypothermia (TH) has been demonstrated to improve survival and neurological outcomes in comatose survivors of out-of-hospital cardiac arrest, in part, by limiting cardiac dysfunction during the immediate post-ROSC period (3, 40). Researchers from our laboratory and others have previously demonstrated that intraarrest hypothermia attenuates cardiac dysf...

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Manipulation of redox signaling in mammalian cells enabled by controlled photogeneration of reactive oxygen species

Cited by 39 publications

References 64 publications

Pulse radiolysis and steady-state analyses of the reaction between hydroethidine and superoxide and other oxidants

Pulse radiolysis and steady-state analyses of the reaction between hydroethidine and superoxide and other oxidants

Beta-Like Importins Mediate the Nuclear Translocation of Mitogen-Activated Protein Kinases

Akt1 genetic deficiency limits hypothermia cardioprotection following murine cardiac arrest

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