Key points are not available for this paper at this time.
The mechanism of senescence-associated cytoplasmic induction of p-Erk1/2 (SA-p-Erk1/2) proteins in human diploid fibroblasts was investigated. p-Erk1/2 proteins were efficiently dephosphorylated in vitro by protein phosphatases 1 and 2A (PP1/2A) and MAPK phosphatase 3 (MKP3). Specific activity of PP1/2A and MKP3 activity significantly decreased during cellular senescence, whereas their protein expression levels did not. To investigate possible mechanism of phosphatase inactivation, we measured reactive oxygen species (ROS) generation by fluorescence-activated cell sorting analysis and found it was much higher in mid-old cells than the young cells. Treating the young cells once with 1 mm H2O2 remarkably induced p-Erk1/2 expression; however, it was transient unless repeatedly treated until 72 h. Multiple treatment of the cells with 0.2 mm H2O2 significantly duplicated inactivation of PP1/2A; however, thiol-specific reagents could reverse the PP1/2A activities, suggesting the oxidation of cysteine molecule in PP1/2A by the increased ROS. When the cells were pretreated with 10 mm N-acetyl-l-cysteine for 1 h, Erk1/2 activation was completely blocked. To elucidate which cysteine residue and/or metal ion in PP1/2A was modified by H2O2, electrospray ionization-tandem mass spectrometry analyses were performed with purified PP1C-α and found Cys62-SO3H and Cys105-SO3H, implicating the tertiary structure perturbation. H2O2 inhibited purified PP1C-α activity by both oxidation of Cys residues and metal ion(s), evidenced by dithiothreitol and ascorbate-restoration assay. In summary, SA-p-Erk1/2 was most likely due to the oxidation of PP1/2A, which resulted from the continuous exposure of the cells to vast amounts of ROS generated during cellular senescence by oxidation of Cys62 and Cys105 in PP1C-α and metal ion(s). The mechanism of senescence-associated cytoplasmic induction of p-Erk1/2 (SA-p-Erk1/2) proteins in human diploid fibroblasts was investigated. p-Erk1/2 proteins were efficiently dephosphorylated in vitro by protein phosphatases 1 and 2A (PP1/2A) and MAPK phosphatase 3 (MKP3). Specific activity of PP1/2A and MKP3 activity significantly decreased during cellular senescence, whereas their protein expression levels did not. To investigate possible mechanism of phosphatase inactivation, we measured reactive oxygen species (ROS) generation by fluorescence-activated cell sorting analysis and found it was much higher in mid-old cells than the young cells. Treating the young cells once with 1 mm H2O2 remarkably induced p-Erk1/2 expression; however, it was transient unless repeatedly treated until 72 h. Multiple treatment of the cells with 0.2 mm H2O2 significantly duplicated inactivation of PP1/2A; however, thiol-specific reagents could reverse the PP1/2A activities, suggesting the oxidation of cysteine molecule in PP1/2A by the increased ROS. When the cells were pretreated with 10 mm N-acetyl-l-cysteine for 1 h, Erk1/2 activation was completely blocked. To elucidate which cysteine residue and/or metal ion in PP1/2A was modified by H2O2, electrospray ionization-tandem mass spectrometry analyses were performed with purified PP1C-α and found Cys62-SO3H and Cys105-SO3H, implicating the tertiary structure perturbation. H2O2 inhibited purified PP1C-α activity by both oxidation of Cys residues and metal ion(s), evidenced by dithiothreitol and ascorbate-restoration assay. In summary, SA-p-Erk1/2 was most likely due to the oxidation of PP1/2A, which resulted from the continuous exposure of the cells to vast amounts of ROS generated during cellular senescence by oxidation of Cys62 and Cys105 in PP1C-α and metal ion(s). Decreased growth rate, limited cell division, flat and large cell shapes, and tight binding of the cells to culture dished (1Macieira-Coelho A. Loria E. Berumen L. Adv. Exp. Med. Biol. 1975; 53: 51-65Crossref PubMed Scopus (22) Google Scholar, 2Start R.D. Loomes R.S. Shortland J.R. Int. J. Exp. Pathol. 1991; 76: 647-654Google Scholar) are well known characteristics of cells entering into replicative senescence (3Hayflick L. Moorehad P.S. Exp. Cell Res. 1961; 25: 585-621Crossref PubMed Scopus (5300) Google Scholar). Primary mouse embryo fibroblasts exposed to oncogenic Ras overexpression undergo premature senescence in response to constitutive MAPK 1The abbreviations used are: MAPK, mitogen-activated protein kinase; PP1, phosphatase 1; and PP2A phosphatase 2A; p-Erk1/2, extracellular signal-regulated protein kinase 1/2; SA-p-Erk1/2, senescence-associated persistent induction of p-Erk1/2; HDF, human diploid fibroblasts; PTP, protein-tyrosine phosphatase; MKPs, MAPK phosphatases; VHR, vaccinia H1-related; rVHR, recombinant VHR; ROS, reactive oxygen species; PBS, phosphate-buffered saline; DTT, dithiothreitol; PMSF, phenylmethylsulfonyl fluoride; pNPP, p-nitrophenyl phosphate; IP, immunoprecipitation; FACS, fluorescence-activated cell sorting; H2-DCFDA, 2′,7′-dichlorodihydrofluorescein diacetate; NAC, N-acetyl-l-cysteine; ESI, electrospray ionization; MS, mass spectrometry; MS/MS, tandem mass spectroscopy; EGF, epidermal growth factor; NBD-Cl, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole; TOF, time-of-flight; bis-Tris, 2-bis(2-hydroxyethyl)amino-2-(hydroxymethyl)propane-1,3-diol.1The abbreviations used are: MAPK, mitogen-activated protein kinase; PP1, phosphatase 1; and PP2A phosphatase 2A; p-Erk1/2, extracellular signal-regulated protein kinase 1/2; SA-p-Erk1/2, senescence-associated persistent induction of p-Erk1/2; HDF, human diploid fibroblasts; PTP, protein-tyrosine phosphatase; MKPs, MAPK phosphatases; VHR, vaccinia H1-related; rVHR, recombinant VHR; ROS, reactive oxygen species; PBS, phosphate-buffered saline; DTT, dithiothreitol; PMSF, phenylmethylsulfonyl fluoride; pNPP, p-nitrophenyl phosphate; IP, immunoprecipitation; FACS, fluorescence-activated cell sorting; H2-DCFDA, 2′,7′-dichlorodihydrofluorescein diacetate; NAC, N-acetyl-l-cysteine; ESI, electrospray ionization; MS, mass spectrometry; MS/MS, tandem mass spectroscopy; EGF, epidermal growth factor; NBD-Cl, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole; TOF, time-of-flight; bis-Tris, 2-bis(2-hydroxyethyl)amino-2-(hydroxymethyl)propane-1,3-diol. kinase/MAPK mitogenic signaling (4Serrano M. Lin A.W. McCurrach M.E. Beach D. Lowe S.W. Cell. 1997; 88: 593-602Abstract Full Text Full Text PDF PubMed Scopus (3861) Google Scholar, 5Lin A.W. Barradas M. Stone J.C. van Aelst L. Serrano M. Lowe S.W. Gene (Amst.). 1998; 12: 3008-3019Google Scholar), whereas established variants lacking p53 or p19ARF are efficiently transformed (6Kamijo T. Zindy R. Roussel M.F. Quelle D.E. Downing J.R. Ashmun R.A. Grosveld G. Sherr C.J. Cell. 1997; 91: 649-659Abstract Full Text Full Text PDF PubMed Scopus (1368) Google Scholar). Ha-Ras mutants with Ras proteins are known and A. A. Aelst L. M. Cell. Full Text PDF PubMed Scopus Google Scholar, T. D. PubMed Scopus Google to and MAPK A. A. Aelst L. M. Cell. Full Text PDF PubMed Scopus Google Scholar), and the with and and cell of T. D. PubMed Scopus Google Scholar, A. D. A. J. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar), the to binding to or A. A. Aelst L. M. Cell. Full Text PDF PubMed Scopus Google Scholar). of the cellular senescence induced by overexpression of the mutants well the replicative senescence in human diploid fibroblasts the increased of extracellular signal-regulated protein kinase G. PubMed Scopus Google Scholar). mechanism MAPK activity by and The activation of the MAPK activity the of the and residues in the kinase activation R. J. Biol. 1991; Full Text PDF PubMed Google Scholar, J. J. 1991; PubMed Scopus Google Scholar, A. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar), and the of protein phosphatase G. T. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar), protein-tyrosine phosphatase G. T. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar, R. A. A. J. 1998; PubMed Scopus Google Scholar), or phosphatases Cell. 1991; Full Text PDF Scopus Google Scholar, J. Biol. Full Text PDF PubMed Google Scholar). The phosphatases are of the of the from well The phosphatases and the p-Erk1/2 are MAPK phosphatases and Cell Biol. 1998; PubMed Scopus Google Scholar, T. T. E. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). MKP3 M. R. M. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar), J. PubMed Scopus Google Scholar) or J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar), in the for Erk1/2 inactivation, and by growth or M. R. M. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar, M. A. M. A. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). of the of MKP3 with Erk1/2 M. A. A. M. A. J. Biol. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar), and phosphatase activity of MKP3 in the M. A. M. 1998; PubMed Scopus Google Scholar). the of MKP3 significantly to VHR, of the J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar), and Erk1/2 proteins are for MKP3 phosphatase to the protein phosphatases to the or protein phosphatases with in cellular of protein 1 2A or and in cells PubMed Scopus Google Scholar, G. PubMed Scopus Google Scholar). in and and and to and which completely activity J. PubMed Scopus Google Scholar). The to both in vitro and in and by protein kinase to of of J. PubMed Scopus Google Scholar, M. E. M. J. Biol. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar, L. A. J. Cell PubMed Google Scholar), whereas of by kinase to activation of the J. PubMed Scopus Google Scholar, J. Biol. Full Text PDF PubMed Google Scholar). PP2A of and PP2A by the inhibited by of PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, A. J. Res. Cell PubMed Scopus Google Scholar). PP2A most in the of whereas and for and PubMed Scopus Google Scholar). are cellular in the to the of growth and A. Biol. Med. PubMed Scopus Google Scholar, L. R. L. Biol. Med. 1997; PubMed Scopus Google Scholar, E. Biol. Med. 1997; PubMed Scopus Google Scholar, A. Biol. Med. 1997; PubMed Scopus Google Scholar, R.A. A. J. 1997; PubMed Scopus Google Scholar, J.C. Full Text PDF PubMed Google Scholar). ROS H2O2 to in growth signaling Google Scholar, M. T. PubMed Scopus Google Scholar), to of H2O2, to transient and inactivation of phosphatase activity by cysteine residue to 1998; PubMed Scopus Google Scholar). levels of H2O2 Erk1/2 J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar, M. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar, A. J. Biol. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar, J. J. Cell Biol. 1998; PubMed Scopus Google Scholar, E. J. 1998; PubMed Scopus Google Scholar) and protein kinase in cells by to D. M. A. D. PubMed Scopus Google Scholar) with to the and and of during with the was in the and of the in the the to the of the mechanism of in M. 1997; PubMed Scopus Google Scholar). in to investigate the mechanism of SA-p-Erk1/2, of PP1/2A and MKP3 with their protein expression levels were during cellular the of ROS generation the young and cells and the of ROS PP1/2A and MKP3 were culture of the and cells. Cell culture of was from the of by the G. PubMed Scopus Google Scholar) and mutants and cell were in by G. PubMed Scopus Google Scholar). cells were in modified with in cells were in the during the cells were to and the of well the were and cells used in the were the with of h, and p-Erk1/2 cells were and with in for the with PBS, cells were with in for and with in for 1 were with in by with PBS, and was with PBS, the was with of and Erk1/2 and cells were with mm mm mm 1 mm 1 mm PMSF, 1 and of cell were in mm protein to for 1 and was the or were was to protein expression In to elucidate the used in both and Erk1/2 proteins or purified and PP2A were to the and cell and analyses were performed well and When the levels of p-Erk1/2 by PP2A in young and which were in the for h, were treated with for and the cells were until for analysis with from was used for phosphatase assay. and were in of and used and of of the and of cell which was in in PBS, 1 mm PMSF, and 1 was in the and the was by in was and phosphatase activity was from the with 1 and 2A the activity of PP1/2A, cells were with and with mm 1 mm PMSF, 1 by for 10 The cell were for 10 and the was used for the assay. To cytoplasmic and cell were with mm mm 10 mm and the was for 10 The was the cytoplasmic and the was in the was by the and for 10 and the was used was with from and of the was for 10 and the was with of The was for for In to from the cell and the the or was for was by the was from the with mm of 1 and 2A by investigate the of ROS in significantly PP1/2A activity during cellular senescence, young cells were pretreated once with 1 mm the PP1/2A activity in the mid-old cells most likely due to H2O2, cells were pretreated with The cells were with 10 and the cell were to the To the H2O2 the cysteine residue in PP1/2A or or was to the young and mid-old cell the PP1/2A assay. of the with young and mid-old cells were 3 the reagents were from of MKP3 MKP3 protein was in the cells by M. R. M. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar) and by MKP3 protein with of was into and of In 3 the was with MKP3 protein with of and the were 1 the to the was and was purified by the with protein MKP3 with was performed with young and cell in mm 1 mm 1 mm 1 mm PMSF, and 1 by the the MKP3 and mm in mm was for 10 h, and the was measured with cell and were for assay. When the of p-Erk1/2 proteins by MKP3 in young and cell MKP3 and the were and analysis was performed p-Erk1/2 of ROS by generation during cellular senescence was measured by analysis 2′,7′-dichlorodihydrofluorescein in cell culture and mid-old cells cells were large to by were in culture with and The were and the cells were in for h. The cells were pretreated with 10 mm for 1 h, and was 10 to cell ROS generation was with by the of from the oxidation of mass was used for analysis of Cys62 and Cys105 were for analysis of once the were the mass was to of the The was used to for in the The was of and the was of 1 was to the in the ion with of to was used for analysis to the of were with and The were of Erk1/2 during young cells with of and the cells with of p-Erk1/2 levels were measured by of p-Erk1/2 was significantly higher in the cells with the young cells analysis response to treatment in the induction of p-Erk1/2 in the young cells and and in the cells and the of p-Erk1/2 was cells To investigate the induction of p-Erk1/2 was limited to replicative senescence premature senescence cells were and analysis was in the constitutive induction of p-Erk1/2 was in both the replicative senescence and oncogenic premature senescence cells and we persistent induction of or increased p-Erk1/2 proteins are in the cells. of and induced premature senescence in cells. were in by and analyses were performed with the cell The of the were the and young and in of p-Erk1/2 phosphatases in the induction of SA-p-Erk1/2 in the of the young and cells were with recombinant phosphatase protein phosphatase or by J. M. and the were by with of phosphatase 10 of PP1C-α of and 0.2 of completely from the p-Erk1/2 of the young cells of the both PP1C-α and the in the cells and SA-p-Erk1/2 by rVHR, by and well the In to well the Erk1/2 proteins or of MAPK was with both and of PP1C-α for 1 h, and the of the protein was by analyses with p-Erk1/2, and Erk1/2 in PP1C-α efficiently dephosphorylated with the Erk1/2 activity p-Erk1/2 proteins was by the purified with cell in mm mm 1 mm PMSF, and for 1 h. When the was by with and Erk1/2 was found to to dephosphorylated by p-Erk1/2 the purified the PP2A could efficiently the of SA-p-Erk1/2, which was increased during cellular SA-p-Erk1/2 and PP2A to well the p-Erk1/2 was Erk1/2 proteins or of MAPK was with or or of PP1C-α for 1 h, and of the protein was by analyses with p-Erk1/2, and Erk1/2 PP1C-α efficiently dephosphorylated it was could the proteins the activity of the purified was with of cell in the of mm mm 1 mm PMSF, and for 1 h. analyses were with p-Erk1/2 and Erk1/2 PP2A dephosphorylated SA-p-Erk1/2 proteins increased in the the of during investigate the of phosphatases were during cellular senescence or phosphatase and PP1/2A were with the of the and cells. in phosphatase activity was and of of protein and PP1/2A were and of of protein in the and cells phosphatases were significantly in the mid-old cells with the young cells. we are with the of Erk1/2 and and their we the of protein phosphatase in during cellular and cytoplasmic were from the young and and possible of with was by in the of the was of both the and cytoplasmic PP1/2A were found to decreased in the than in the young cells The the of PP1/2A activity in the cells was due to expression of was by analysis with and however, the young and cells was in to the of PP2A the SA-p-Erk1/2 in the young and cells were treated with PP2A PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, A. J. Res. Cell PubMed Scopus Google Scholar). the cells were and the and cell were by in of PP2A with significantly increased levels of Erk1/2 proteins in both the young and the of and PP2A the SA-p-Erk1/2 in both young and cells. In to the PP1/2A proteins expression and their in the young and PP1/2A activity and of protein in cells were in the cells protein and higher PP1/2A activity well much cell than the young cells. the activity in the cells was of the young activity of PP1/2A was significantly decreased during cellular of protein phosphatase 1 and 2A in young and cells MKP3 for of p-Erk1/2 p-Erk1/2 to to the of MKP3 phosphatase J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar), and in the of M. R. M. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar), the of MKP3 to SA-p-Erk1/2 in the was by the amounts of p-Erk1/2 to MKP3 in the the and the by p-Erk1/2 in large of p-Erk1/2 protein was to MKP3 in the whereas was p-Erk1/2 to MKP3 in the young cells. we measured the MKP3 protein expression in both the young and cells with analysis was during the activity was measured by the activity in the cells was significantly than in the young cells the of of PP1/2A and MKP3 during to investigate the mechanism the of PP1/2A and MKP3 and p-Erk1/2 during cellular senescence, ROS generation was measured during cellular senescence by analysis both the young and mid-old cells with in the mid-old cells ROS than the young and the ROS generation by both could efficiently inhibited by with 10 mm When young cells were treated once with 1 mm H2O2, Erk1/2 was increased 10 however, the was in the the cells were pretreated for 1 with 10 mm NAC, of Erk1/2 protein was efficiently inhibited To H2O2 was for the of PP1/2A or the were measured treatment of the young cells once with in the were inhibited in 10 to and to the in h. To investigate the of p-Erk1/2 and activity of PP1/2A by H2O2 we measured the of the by analysis and the activity in the young cells were repeatedly treated with H2O2 h, PP1/2A significantly inhibited 72 young cells activity of and it to and protein and h, with the of PP1/2A activities, of Erk1/2 proteins was by H2O2 of the young cells by H2O2 exposure inhibited PP1/2A activities, in the induction of p-Erk1/2 To elucidate of PP1/2A in the mid-old cells was due to the oxidation of cysteine residues in the or thiol-specific DTT, and were to the of the young which pretreated with H2O2 for 10 in the of the PP1/2A activity was decreased from to protein by the treatment with the was to and protein by the of DTT, and NAC, reagents significantly PP1/2A activity with of the H2O2 treatment and DTT, and NAC, significantly increased PP1/2A activity of the young cells than the the was with mid-old cell PP1/2A activity most likely due to H2O2 during senescence, could significantly PP1/2A activity with the in suggesting the of ROS found in cells which are for phosphatase To investigate of ROS generated during cellular senescence could the to cells or the mid-old cells were treated with for and and their was in the flat and large cell to the to the to young and In to investigate the of the cells were by to cellular senescence, the expression of senescence-associated in the cells was in treatment of the cells with for was to significantly senescence-associated expression in the mid-old of mid-old cells to cells by cells were treated with for in of the mid-old cells to young of expression by of mid-old cells with in and/or in PP1C-α by oxidation of by H2O2 in mm was with purified PP1C-α and 1998; PubMed Scopus Google Scholar). in of PP1C-α with mm H2O2 for 10 to by and the of from to To elucidate the of their activities, and were to PP1C-α with mm H2O2 for 10 of PP1C-α could by by To the most of oxidation in cysteine of the purified analysis was performed mass in and were in the of and The are with the oxidation of Cys62 to Cys62-SO3H and Cys105 to the of mass In oxidation of were in for Cys62 mass of When H2O2 PP1C-α with the oxidation whereas and were To the of the were to The of the ion from and and were from the of the from of the and it was to in the from to to Cys62-SO3H and and the mass and of spectrometry analysis of oxidation of and PP1C-α were with for and analysis by MS/MS, were with of into and the and and are tandem mass of with and The senescence of human fibroblasts used in vitro PubMed Scopus Google Scholar). In the we large of p-Erk1/2 SA-p-Erk1/2, were found in the of both and cells induced by mutants 1 and and the were for PP1/2A, PTP, and in vitro well in the of cellular senescence, PP1/2A and MKP3 were significantly however, their protein expression levels were and The to with the the phosphatase were in the mid-old cells and large of SA-p-Erk1/2 proteins was and by MKP3 in the of cells Erk1/2 of MAPK with protein kinase and and the of Erk1/2 and MAPK kinase R. J. PubMed Scopus Google Scholar, R. C.J. Google Scholar). When cells are with growth or Erk1/2 the p-Erk1/2 into the and for cell R. A. J. 12: PubMed Scopus Google Scholar). The of protein in the the of phosphatases and and or protein phosphatases with in cellular the of in both the and of the cells in their protein expression resulted in the induction of p-Erk1/2 levels by the and induction of phosphatase or during cellular in to the or of in the we of the known 1 expression in the young and cells by and the to the PP1/2A activity the young and cell When by the protein expression levels the were the amounts of the cell were to of young cells and the PP1/2A activity was the of the activity measured were the in the the or of in the cell The the the decreased PP1/2A activity in the cells was due to of in the cells. To investigate the of the PP1/2A activity during cellular senescence, thiol-specific were to the cell cells pretreated with in the could significantly the PP1/2A in the young cells and the PP1/2A activity in mid-old cells. the of phosphatase activity due to the oxidation of reactive cysteine by ROS during cellular the of PP1, and are and PP2A and PP2A and their and with are The structure of their of which J. J. PubMed Scopus Google Scholar, A. R. PubMed Scopus Google Scholar). the of the activity of D. PubMed Scopus Google Scholar) the of PP1/2A in cell were inhibited by activity could by cells with or with the to the mechanism of of PP1/2A in cells was due to of H2O2 cysteine of PP1/2A due to in 3 and SA-p-Erk1/2 for the of and PP2A by are and A. G. M. T. J. PubMed Scopus Google Scholar), induction of p-Erk1/2 in the young and cells the treatment with and the inactivation of PP2A during cellular senescence could significantly for SA-p-Erk1/2 The induction of p-Erk1/2 by treatment with 10 mm H2O2 was in found in the induction of MAPK M. J. E. M. Biol. PubMed Scopus Google Scholar, T. Biol. Med. PubMed Scopus Google Scholar), and the transient of and PP2A in which were significantly inhibited in 10 by and the in to the cells were treated with H2O2 for than 72 h, of PP1/2A were and the of p-Erk1/2 was in cells in exposed to the higher of ROS during the senescence in the of PP1/2A and the constitutive of was by the ROS measured in the mid-old cells than the young cells When analysis was to ROS from the cells was to the was large and of the of the the of the mid-old cells for the the cells were with 1 mm H2O2 and induced p-Erk1/2 the induction could completely by treatment of the young cells once with of H2O2 significantly inhibited phosphatase activity with the of p-Erk1/2 proteins the treated repeatedly with H2O2 h, the PP1/2A significantly inhibited 72 The levels of p-Erk1/2 in the young cells were with the PP1/2A by the treatment of the mid-old cells with induced of the to the young cells with the of senescence-associated expression H2O2 could by of species H2O2 activity by cysteine oxidation by oxidation of metal which were in the of the PP1C-α D. J. Biol. PubMed Scopus Google Scholar). was evidenced by of activity much the oxidation of metal in the during cellular The of ROS was higher in the mid-old cells than the young the cysteine residue by H2O2 was by mm H2O2 was of H2O2 completely Cys62 and Cys105 of PP1C-α to and and however, the of oxidation could during the to A. J. Biol. Full Text PDF PubMed Google Scholar). oxidation of Cys62 for the PP1/2A activity during cellular senescence, of the The of tertiary structure the of PP1C-α H2O2 generated during cellular senescence both PP1/2A and p-Erk1/2 oxidation of cysteine residues of PP1/2A, suggesting H2O2 the of the significantly PP1/2A activity in the cells. the of activity in of the was significantly the and the was the activity in the which the in and human D. M. A. D. PubMed Scopus Google Scholar), the of PP1/2A and of Erk1/2 proteins in the mid-old and cells in the of PP1/2A during cellular Biol. Cell. PubMed Scopus Google Scholar) 1 protein with the the and of are in cells by the of their cells amounts of the and increased of ROS the of the for most the and the and of into the M.E. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). binding to of the for of with M. T. Cell Biol. PubMed Scopus Google Scholar). of into the higher H2O2 in the mid-old cells than in the young cells MKP3 with Erk1/2 M. A. A. M. A. J. Biol. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar), and the phosphatase activity of MKP3 M. A. M. 1998; PubMed Scopus Google Scholar) of the MKP3 in the and J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). In the the the from the and the by the and residues are for the binding of to the of MKP3 the of residues and the by the activation of MKP3 J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). The of MKP3 and for binding J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). the for MKP3 with of of higher than the L. J. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). H2O2 found in the mid-old cells was most likely for inactivation of MKP3 during cellular of the for the reactive and H2O2 to Cys to 1998; PubMed Scopus Google Scholar), it was possible H2O2 the residue in MKP3 during senescence, in the of of the MKP3 large of p-Erk1/2 in the cells it to p-Erk1/2 and the proteins in In summary, SA-p-Erk1/2 proteins in the of cells were due to the PP1/2A and inactivation of cytoplasmic in of p-Erk1/2 in the and of their to the of of p-Erk1/2 in or of in cells. are most by oxidation of the Cys62 in PP1/2A and the the of MKP3 by ROS, which was generated during the senescence for and for the for The and was the of from the
Kim et al. (Mon,) studied this question.