Krüppel-like factor 8 is a novel androgen receptor co-activator in human prostate cancer
Abstract
Hong-jiang HE1, Xue-feng GU3, Wan-hai XU2, De-jun YANG2, Xiao-min WANG2, Yu SU2, *
1Department of Otolaryngology, Head and Neck Surgery, the Third Affiliated Hospital of Harbin Medical University, Harbin 150081, China; 2Department of Urinary Surgery, the Fourth Affiliated Hospital of Harbin Medical University, Harbin 150001, China; 3Clinical Research Center, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, China
Aim: Krüppel-like factor 8 (KLF8) plays important roles in cell cycle and oncogenic transformation. On other hand, androgen receptor (AR) is crucial in development of both androgen-dependent and independent prostatic malignancies. The aim of this study is to investigate the role of KLF8 in prostate cancer (PCa) and the relationship between KLF8 and AR.
Methods: Eight human PCa cell lines, including androgen-dependent LNCap cells and androgen-independent 22Rv1 cells, as well as human PCa samples were studied. LNCap cells and 22Rv1 cells were transfected with plasmids encoding full-length wild-type KLF8 or KLF8 shRNA. The expression of KLF8 protein was detected using Western blotting or immunohistochemical staining. Cell proliferation in vitro was measured with MTT assay, and in vivo in a xenograft nude mouse model. Yeast two-hybrid screening, co-immunoprecipitation and pull down assays were used to examine the binding of KLF8 to AR. Luciferase reporter gene assay was used to measure the transcriptional activity of the genes targeted by AR.
Results: In 133 human PCa samples, KLF8 protein staining was observed in 92.65% (63/68) of high-grade PCa, 66.15% (43/65) of low-grade PCa, and 6.82% (3/44) of adjacent normal tissues. The expression of KLF8 was significantly associated with poorer overall survival. Overexpression of KLF8 enhanced the proliferation of both LNCap and 22Rv1 cells, while knockdown of endogenous KLF8 suppressed the proliferation. These manipulations exerted similar effects on the tumor volumes in the xenograft nude mouse model. Yeast two-hybrid screening revealed that KLF8 was a novel AR-interacting protein. With pull down assay and co-immunoprecipitation assay, we demonstrated that KLF8 bound directly to AR, and KLF8 enhanced AR target gene transcription.
Conclusion: The results demonstrate that KLF8 is a novel AR transcriptional co-activator that is overexpressed in PCa and may play a role in progression of hormone-refractory PCa.
Keywords: Krüppel-like factor 8 (KLF8); prostate cancer; Gleason score; PSA value; androgen receptor; transcriptional co-activator
This study was supported by the Heilongjiang Province Health Department Fund (No 2010-135).
* To whom correspondence should be addressed.
E-mail yusufahhmu@gmail.com
Received 2012-07-02 Accepted 2012-08-16
Keywords:
1Department of Otolaryngology, Head and Neck Surgery, the Third Affiliated Hospital of Harbin Medical University, Harbin 150081, China; 2Department of Urinary Surgery, the Fourth Affiliated Hospital of Harbin Medical University, Harbin 150001, China; 3Clinical Research Center, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, China
Aim: Krüppel-like factor 8 (KLF8) plays important roles in cell cycle and oncogenic transformation. On other hand, androgen receptor (AR) is crucial in development of both androgen-dependent and independent prostatic malignancies. The aim of this study is to investigate the role of KLF8 in prostate cancer (PCa) and the relationship between KLF8 and AR.
Methods: Eight human PCa cell lines, including androgen-dependent LNCap cells and androgen-independent 22Rv1 cells, as well as human PCa samples were studied. LNCap cells and 22Rv1 cells were transfected with plasmids encoding full-length wild-type KLF8 or KLF8 shRNA. The expression of KLF8 protein was detected using Western blotting or immunohistochemical staining. Cell proliferation in vitro was measured with MTT assay, and in vivo in a xenograft nude mouse model. Yeast two-hybrid screening, co-immunoprecipitation and pull down assays were used to examine the binding of KLF8 to AR. Luciferase reporter gene assay was used to measure the transcriptional activity of the genes targeted by AR.
Results: In 133 human PCa samples, KLF8 protein staining was observed in 92.65% (63/68) of high-grade PCa, 66.15% (43/65) of low-grade PCa, and 6.82% (3/44) of adjacent normal tissues. The expression of KLF8 was significantly associated with poorer overall survival. Overexpression of KLF8 enhanced the proliferation of both LNCap and 22Rv1 cells, while knockdown of endogenous KLF8 suppressed the proliferation. These manipulations exerted similar effects on the tumor volumes in the xenograft nude mouse model. Yeast two-hybrid screening revealed that KLF8 was a novel AR-interacting protein. With pull down assay and co-immunoprecipitation assay, we demonstrated that KLF8 bound directly to AR, and KLF8 enhanced AR target gene transcription.
Conclusion: The results demonstrate that KLF8 is a novel AR transcriptional co-activator that is overexpressed in PCa and may play a role in progression of hormone-refractory PCa.
Keywords: Krüppel-like factor 8 (KLF8); prostate cancer; Gleason score; PSA value; androgen receptor; transcriptional co-activator
This study was supported by the Heilongjiang Province Health Department Fund (No 2010-135).
* To whom correspondence should be addressed.
E-mail yusufahhmu@gmail.com
Received 2012-07-02 Accepted 2012-08-16