Identification of XAF1 as a novel cell cycle regulator through modulating G(2)/M checkpoint and interaction with checkpoint kinase 1 in gastrointestinal cancer.
OBJECTIVE
X-linked inhibitor of apoptosis-associated factor 1 (XAF1) was first recognized as an antagonist of X-linked inhibitor of apoptosis in suppressing caspase 3 activity. It has lower expression in cancer cells than normal tissue. Overexpression of XAF1 can inhibit cancer cell growth and sensitize tumor necrosis factor-related apoptosis-inducing ligand- or etoposide-induced apoptosis. The aim of this study is to elucidate the mechanism of XAF1 in regulating cell growth.


METHODS
Stable transfectants of gastrointestinal (GI) cancer cell lines AGS and SW1116 expressing XAF1 and vector control were generated. Cell growth, apoptosis, mitotic status and cell cycle distribution were assessed. The interaction between XAF1 and G(2)/M checkpoint proteins was evaluated by immunoblotting, kinase assay and co-immunoprecipitation assay. Mitotic catastrophe was identified by occurrence of aberrant nuclei and centrosomal amplification.


RESULTS
Our results showed that overexpression of XAF1 suppressed serum-dependent cancer cell growth, induced mitotic catastrophe and G(2)/M cell cycle arrest. Interestingly, XAF1 was predominantly expressed in G(2)/M phase after cell cycle synchronization. XAF1 interacted with and activated checkpoint kinase 1 (Chk1), inactivated Cdc25C and lead to inactivation of Cdc2-cyclin B complex. Suppression of Chk1 abrogated XAF1-induced G(2)/M arrest.


CONCLUSIONS
Our findings implicate XAF1 as a novel cell cycle modulator that is recruited in G(2)/M phase and thus unravel a novel function pathway of XAF1, suggesting the potential role of XAF1 as the target for the management of GI cancers.
Wang
Jide
J
Department of Digestive Medicine, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China.
Gu
Qing
Q
Li
Ming
M
Zhang
Wenjing
W
Yang
Mo
M
Zou
Bing
B
Chan
Shing
S
Qiao
Liang
L
Jiang
Bo
B
Tu
Shuiping
S
Ma
Juan
J
Hung
Ivan F
IF
Lan
Hui Yao
HY
Wong
Benjamin C Y
BC
D048868:Adaptor Proteins, Signal Transducing; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D016203:CDC2 Protein Kinase; D053148:Caspase 3 / Q000378:metabolism; D002453:Cell Cycle; D002455:Cell Division; D002460:Cell Line; D049109:Cell Proliferation; D000071877:Checkpoint Kinase 1; D019926:Cyclin B / Q000502:physiology; D018844:Cyclin-Dependent Kinases; D004249:DNA Damage; D016195:G2 Phase; D005770:Gastrointestinal Neoplasms / Q000473:pathology*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008938:Mitosis; D009363:Neoplasm Proteins / Q000502:physiology*; D011494:Protein Kinases / Q000502:physiology*
D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D019926:Cyclin B; D047908:Intracellular Signaling Peptides and Proteins; D009363:Neoplasm Proteins; C417551:XAF1 protein, human; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D016203:CDC2 Protein Kinase; C528899:CDK1 protein, human; D018844:Cyclin-Dependent Kinases; D053148:Caspase 3