T-LAK cell-originated protein kinase (TOPK) phosphorylation of Prx1 at Ser-32 prevents UVB-induced apoptosis in RPMI7951 melanoma cells through the regulation of Prx1 peroxidase activity.
Protein kinases are potential targets for the prevention and control of UV-induced skin cancer. T-cell-originated protein kinase (TOPK) is highly expressed in skin cancer cells, but its specific function is still unknown. We investigated the role of TOPK in UVB-induced apoptosis in RPMI7951 human melanoma cells. Liquid chromatography-tandem mass spectrometry analysis was used to identify proteins that bind with TOPK. Immunofluorescence, Western blot, and flow cytometry were used to assess the effect of UVB on TOPK, peroxiredoxin 1 (Prx1), and apoptosis in RPMI7951 cells. TOPK binds with Prx1 and its phosphorylation of Prx1 at Ser-32 is important for regulation of H(2)O(2)-mediated signal transduction. Analysis of the CD spectra of Prx1 and mutant Prx1 (S32A) proteins showed that the secondary structure of Prx1 was significantly altered by phosphorylation of Prx1 at Ser-32. UVB irradiation induced phosphorylation of TOPK in RPMI7951 human melanoma cells and phosphorylated TOPK co-localized with Prx1 in the nucleus. UVB induced the peroxidase activity of Prx1 in vitro and ex vivo. Following treatment with UVB, H(2)O(2) levels and apoptosis were increased in RPMI7951 cells stably expressing TOPK siRNA or stably mutant Prx1 (S32A). Phosphorylation of Prx1 (Ser-32) by TOPK prevents UVB-induced apoptosis in RPMI7951 melanoma cells through regulation of Prx1 peroxidase activity and blockade of intracellular H(2)O(2) accumulation.
Zykova
Tatyana A
TA
Hormel Institute, University of Minnesota, Austin, Minnesota 55912, USA.
Zhu
Feng
F
Vakorina
Tatyana I
TI
Zhang
Jishuai
J
Higgins
Lee Ann
LA
Urusova
Darya V
DV
Bode
Ann M
AM
Dong
Zigang
Z
D017209:Apoptosis / Q000502:physiology / Q000528:radiation effects*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002942:Circular Dichroism; D005434:Flow Cytometry; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D047468:Immunoprecipitation; D066298:In Vitro Techniques; D008545:Melanoma / Q000201:enzymology*; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D020929:Mitogen-Activated Protein Kinase Kinases; D054464:Peroxiredoxins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering; D012694:Serine / Q000737:chemistry / Q000378:metabolism*; D014466:Ultraviolet Rays / Q000009:adverse effects*
D034741:RNA, Small Interfering; D012694:Serine; D006861:Hydrogen Peroxide; D054464:Peroxiredoxins; D017346:Protein Serine-Threonine Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; C409022:PDZ-binding kinase