New Modes of GPCR Signalling

Cyclin D1 Promotes Anchorage-independent Cell Survival by Inhibiting
FOXO-Mediated Anoikis
Lu Gan 1, Xiangliang Yang 1, Huibi Xu 1, Haojie Huang 2
1. College of Life Science and Technology, Huazhong University of Science and
Technology, Wuhan 430074, China
2. Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA
Aim: Adhesion to an appropriate extracellular matrix (ECM) is important for normal
cells to survive, and detachment from such supportive matrices usually triggers a
specific type of apoptosis termed anoikis. However, most cancer cells are commonly
resistant to anoikis and can survive in the presence of an inappropriate ECM or in the
absence of anchorage to the ECM. O-class Forkhead box (FOXO) transcription factors
are critical regulators of diverse cellular processes, including apoptosis, cell-cycle arrest,
DNA damage repair and oxidative stress resistance. In this study we determined
whether FOXOs are involved in anoikis and its mechanism. Methods: Annexin V-PE
apoptosis detection kit and soft agar colony formation assay were used to measure the
anoikis in prostate cancer cells. FOXOs activities were determined using a generic
FOXO luciferase reporter gene that contains three copies of forkhead response elements.
Immunoprecipitation was used to determine the endogenous and exogenous interaction
between cyclin D1 and FOXO. In vitro protein binding assay was used to confirm the
interaction between the two proteins. In vitro kinase activity determined whether
FOXO1 was phosphorylated by CDK4/CDK6. Results: FOXO1 and FOXO3a play an
essential role in promoting cell detachment-induced anoikis. In contrast, elevated cyclin
D1 inhibits anoikis. We further show that cyclin D1 interacts with FOXO proteins and
impedes their transcriptional regulatory and anoikis-promoting functions. This effect of
cyclin D1 requires its transcription repression domain but is independent of
cyclin-dependent kinases CDK4 and CDK6. Moreover, we demonstrate that
cancer-derived mutants of cyclin D1 are much more stable than wild-type cyclin D1
under anchorage-independent conditions and possess a greater antagonistic effect on
FOXO-regulated anoikis and anchorage-independent growth of cancer cells.
Conculsion: These data suggest that cyclin D1 may play a critical role in tumorigenesis
and cancer metastasis by inhibiting the anoikis-promoting function of FOXO proteins.
Keywords: Foxo; cyclin D1; Anoikis; protein-protein interaction
Author: Lu Gan,PhD, College of Life Science and Technology, Huazhong University
of Science and Technology, Wuhan 430074, China, Tel: 86-27-87792147
E-mail: lugan@mail.hust.edu.cn