Phosphorylation of CLK2 at Serine 34 and Threonine 127 by AKT Controls Cell Survival after Ionizing Radiation

Seon Young Nam,Cha Soon Kim,Young-Woo Jin,Meeseon Jeong,Hyung Ho Seo,Sungkwan An,Kwang Hee Yang,Hyung Sun Park,Ji-Young Kim

doi:10.1074/jbc.m110.122044

Abstract

AKT phosphorylates components of the intrinsic cell survival machinery and promotes survival to various stimuli. In the present study, we identified CDC-like kinase 2 (CLK2) as a new substrate of AKT activation and elucidated its role in cell survival to ionizing radiation. AKT directly binds to and phosphorylates CLK2 on serine 34 and threonine 127, in vitro and in vivo. CLK2 phosphorylation was detected in HeLa cells overexpressing active AKT. In addition, we demonstrated that ionizing radiation induces CLK2 phosphorylation via AKT activation. In contrast, the suppression of endogenous AKT expression by siRNA inhibited CLK2 phosphorylation in response to 2 gray of γ-ray or insulin. Furthermore, we examined the effect of CLK2 on the survival of irradiated CCD-18Lu cells overexpressing Myc-CLK2. CLK2 overexpression significantly increased cell growth and inhibited cell death induced by 2 gray. The role of CLK2 in cell survival to ionizing radiation was dependent on the phosphorylation of serine 34 and threonine 127. Our results suggest that AKT activation controls cell survival to ionizing radiation by phosphorylating CLK2, revealing an important regulatory mechanism required for promoting cell survival.

Highlights

Ionizing radiation induces a variety of cellular responses, such as DNA damage and repair, cell cycle arrest, apoptosis, and carcinogenesis (10 –15)
To eliminate the possibility of the potential off-target effect by specific for AKT (siAKT), we examined whether AKT activation is correlated with the regulation of p-MEK1/2 and MEK1/2 expression, which is an upstream molecule of mitogen-activated protein kinases (MAPKs)
We identified and characterized CDC-like kinase 2 (CLK2) as a novel substrate of AKT

Summary

Introduction

Ionizing radiation induces a variety of cellular responses, such as DNA damage and repair, cell cycle arrest, apoptosis, and carcinogenesis (10 –15). AKT directly phosphorylates acinus and regulates its expression via the nuclear factor ␬B pathway, thereby controlling cell survival to ionizing radiation (20, 21). We first show that CLK2 is a novel substrate for AKT and that CLK2 phosphorylation by activated AKT leads to cell survival following ionizing radiation.

Results

Conclusion