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

Tatyana A Zykova,Feng Zhu,Tatyana I Vakorina,Jishuai Zhang,Lee Ann Higgins,Darya V Urusova,Ann M Bode,Zigang Dong

doi:10.1074/jbc.m110.135905

Tatyana A Zykova, Feng Zhu + Show 6 more

Open Access

https://doi.org/10.1074/jbc.m110.135905

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Abstract

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.

Highlights

Its kinase activity [6]
Our results demonstrate that T-cell-originated protein kinase (TOPK) phosphorylation of peroxiredoxin 1 (Prx1) at Ser-32 inhibits UVB-induced apoptosis in RPMI7951 melanoma cells by increasing Prx1 peroxidase activity and decreasing the intracellular accumulation of H2O2
We found that TOPK was more abundant in malignant melanoma tissues compared with squamous and basal carcinoma tissues and that UVB increased the phosphorylation of TOPK in RPMI7951 melanoma cells

Summary

Introduction

Its kinase activity [6]. TOPK expression is regulated by cell cycle-specific transcription factors E2F and CREB/ATF [7]. TOPK is found in activated T-LAK cells, lymphoid tumor cells, and normal testicular tissue [1]. Earlier we showed that TOPK is involved in preventing apoptosis in melanoma cells [13] and is a positive regulator of c-Jun-NH2-kinase 1 (JNK1) signaling and H-Rasinduced cell transformation [15]. Prx plays a negative role in apoptosis signal-regulating kinase 1 (ASK1)-induced apoptosis [21], and its overexpression in cancer cells could enhance survival. Our results demonstrate that TOPK phosphorylation of Prx at Ser-32 inhibits UVB-induced apoptosis in RPMI7951 melanoma cells by increasing Prx peroxidase activity and decreasing the intracellular accumulation of H2O2

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