title: [The role of protein phosphatase 2A B56β holoenzyme in the regulation of heavy metal CdCl2 induced cytotoxicity].
abstract: OBJECTIVE
  To investigate the role of holoenzyme containing Protein Phosphatase 2A B56β in regulating CdCl2 induced cytotoxicity.
  
  
  METHODS
  CdCl2-induced cytotoxicity in normal human cell line L-02, AFB1-transformed hepatic cell line L-02 RT-AFB1 and tumor cell line Bel7402 was measured by modified MTT assay. Stable cell lines L-02 SHAKT, L-02 SHB56β, L-02 RT-AFB1-B56β and Bel7402-B56β were generated by infecting L-02 cells or Bel7402 cells with retroviral vectors encoding lentiviral AKT shRNA, lentiviral B56β shRNA and B56β. The relative cell viability was measured in normal human cell line AFB1-transformed hepatic cell line and tumor cell line when treated by CdCl2 (0, 20, 40, 80, 160 µmol/L). After treated by wortmannin (2.5, 5.0 µmol/L) combined with 40 µmol/L CdCl2, Western blot was applied to measure the expression of associated protein in L-02.Western blot was applied to measure the expression of B56β, MT (metallothionein), AKT, and p-AKT in these cell lines treated by CdCl2.
  
  
  RESULTS
  The levels of MT were 0.12 ± 0.02, 0.06 ± 0.06 in L-02 RT-AFB1 and Bel7402, which were lower than L02 (0.92 ± 0.14) (F = 1 148.16, P < 0.001) when treated by 40 µmol/L CdCl2. When treated by 40 µmol/L CdCl2, the expression of p-AKT in L-02 SHAKT-1 and L-02 SHAKT-2 were 0.08 ± 0.02, 0.08 ± 0.05, which levels were lower than L-02 SHGFP (0.18 ± 0.15) (F = 724.70, P < 0.001); and the expression of MT were both 0.62 ± 0.16 in L-02 SHAKT-1 and L-02 SHAKT-2, which levels were higher than L-02 SHGFP (0.22 ± 0.14) (F = 94.73, P < 0.001). After treated by wortmannin (2.5, 5.0 µmol/L) combined with 40 µmol/L CdCl2, the expression of p-AKT in L-02 were 0.28 ± 0.07, 0.15 ± 0.11, which levels were lower than wortmannin untreated cells (0.52 ± 0.11) (F = 578.57, P < 0.001); and the expreesion of MT were 1.62 ± 0.80, 1.08 ± 0.15, which levels were higher than wortmannin untreated cells (0.69 ± 0.18) (F = 12.34, P < 0.001). When treated by 40 µmol/L CdCl2, the levels of p-AKT in L-02 SHB56β-1 and L-02 SHB56β-2 were 0.57 ± 0.13, 0.59 ± 0.02, which were higher than L-02 SHGFP (0.32 ± 0.02) (F = 87.16, P < 0.001); and the levels of MT were 0.35 ± 0.07, 0.20 ± 0.03 in L-02 SHB56β-1 and L-02 SHB56β-2, which were lower than L-02 SHGFP (1.51 ± 0.13) (F = 2 457.10, P < 0.001). After treated by 40 µmol/L CdCl2, the expression of p-AKT in L-02 RT-AFB1-B56β and Bel7402-B56β were 0.10 ± 0.11, 0.09 ± 0.01, which were lower than L-02 RT-AFB1 (0.36 ± 0.01) and Bel7402 (0.43 ± 0.11) (F = 877.62, P < 0.001); and the levels of MT were 0.92 ± 0.13, 0.95 ± 0.08 in L-02 RT-AFB1-B56β and Bel7402-B56β,which were higher than L-02 RT-AFB1 (0.44 ± 0.12) and Bel7402 (0.77 ± 0.06) (F = 51.97, P < 0.001).
  
  
  CONCLUSIONS
  Protein phosphatase 2A complexes containing B56β participated in the regulation of MT expression through direct dephosphorylation of AKT, finally affected the cytotoxicity responding to CdCl2. Our study revealed a key signaling pathways of PP2A involved in heavy metals induced cytotoxicity.
authors.affiliation: Faculty of Preventive Medicine, School of Public Health, Sun-Yat Sen University, Guangzhou 510080, China.
authors.forename: Jinmiao
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Faculty of Preventive Medicine, School of Public Health, Sun-Yat Sen University, Guangzhou 510080, China.
authors.forename: Lu
authors.initials: L
authors.lastname: Ma
authors.affiliation: Faculty of Preventive Medicine, School of Public Health, Sun-Yat Sen University, Guangzhou 510080, China.
authors.forename: San
authors.initials: S
authors.lastname: Wang
authors.affiliation: Faculty of Preventive Medicine, School of Public Health, Sun-Yat Sen University, Guangzhou 510080, China.
authors.forename: Wen
authors.initials: W
authors.lastname: Chen
authors.affiliation: Faculty of Preventive Medicine, School of Public Health, Sun-Yat Sen University, Guangzhou 510080, China; Email: chenliping_happy06@163.com.
authors.forename: Liping
authors.initials: L
authors.lastname: Chen
mesh_terms: D019256:Cadmium Chloride; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival; D020035:Holoenzymes; D006801:Humans; D008099:Liver; D008668:Metallothionein; D019216:Metals, Heavy; D054648:Protein Phosphatase 2; D015398:Signal Transduction
chemical_list: D020035:Holoenzymes; D019216:Metals, Heavy; D008668:Metallothionein; D054648:Protein Phosphatase 2; D019256:Cadmium Chloride