Lymphokine-Activated Killer T-Cell-Originated Protein Kinase Phosphorylation of Histone H2AX Prevents Arsenite-Induced Apoptosis in RPMI7951 Melanoma Cells

Abstract

Purpose: Arsenic is a valuable therapeutic tool in cancer treatment. Lymphokine-activated killer T-cell-originated protein kinase (TOPK) is highly expressed in cancer cells, but its specific function is still unknown. We investigated the role of TOPK in arsenic-induced apoptosis in RPMI7951human melanoma cells. Experimental Design: Expression of TOPK was evaluated in different melanoma cell lines, and liquid chromatography-tandem mass spectrometry analysis was used to identify proteins binding with TOPK. Immunofluorescence, Western blot, and flow cytometry were used to assess the effect of arsenic onTOPK, histone H2AX, and apoptosis in RPMI7951cells. Results: Melanoma cell lines expressing high levels of TOPK were more resistant to arsenite (As 3+ )-induced apoptosis. As 3+ treatment induced phosphorylation of TOPK and histone H2AX in RPMI7951human melanoma cells. Liquid chromatography-tandem mass spectrometry results indicated thatTOPK could bind with histone H2AX, and in vitro and in vivo assays confirmed that TOPK binds with and phosphorylates histone H2AX. As 3+ treatment caused phosphorylation of TOPK, which colocalized with phosphorylated histone H2AX in the nucleus. TOPK small interfering RNA cells exhibited a decreased phosphorylation of histone H2AX with As 3+ treatment. As 3+ -induced apoptosis was decreased in H2AX À/À cells but increased in TOPK small interfering RNA cells. Conclusions: TOPK binds with histone H2AX and inhibits As 3+ -induced apoptosis through phosphorylation of histone H2AX. Melanoma cell lines with high levels ofTOPK are more resistant to As 3+ -induced apoptosis. Therefore, inhibition of TOPK activity combined with As 3+ treatment may be helpful in the treatment of melanomas.

PBK/TOPK interacts with the DBD domain of tumor suppressor p53 and modulates expression of transcriptional targets including p21

“…Cellcycle-specific regulation of PBK/TOPK is mediated partly by transcription factors E2F and CREB/ATF (Nandi and Rapoport, 2006). Previous studies also suggest a role for PBK/TOPK in cytokinesis and DNA damage sensing and repair through phosphorylation of histone H2AX (Matsumoto et al, 2004;Zykova et al, 2006;Ayllo´n and O'Connor, 2007).…”

TOPK/PBK promotes cell migration via modulation of the PI3K/PTEN/AKT pathway and is associated with poor prognosis in lung cancer

“…TOPK is a 322 amino-acid MAPKK-like serine/threonine kinase and is highly expressed in various types of cancer, such as lymphoma, leukemia, breast cancer, melanoma and colorectal cancers, but is undetectable in normal tissues except the germ cells of testis and several fetal tissues (Abe et al, 2000;Simons-Evelyn et al, 2001;Nandi et al, 2004;Park et al, 2006;Zykova et al, 2006;Zhu et al, 2007). In cultured cells, the expression of TOPK is upregulated during mitosis, when it is thought to be phosphorylated on Thr 9 and activated by cyclin B1/cdk1 (Matsumoto et al, 2004).…”

Phosphorylation of H2AX at Ser139 and a New Phosphorylation Site Ser16 by RSK2 Decreases H2AX Ubiquitination and Inhibits Cell Transformation

“…We reported previously that ultraviolet A (UVA) irradiation induces histone H2AX phosphorylation that is mediated by c-Jun N-terminal kinases (JNK), and the JNKs/ H2AX pathway is associated with the induction of apoptosis (12). We also found that T-cell-originated protein kinase (TOPK) directly phosphorylates histone H2AX and is involved in arsenic-induced apoptosis of melanoma cells (13). Histone H2AX phosphorylation is induced by various stresses, such as replication stress (9,12,14,15), endogenous stress (16), ionizing radiation (17), and other agents that cause DNA damage (18), including DNA DSBs (19,20).…”