Nucleoporin 62 and Ca(2+)/calmodulin dependent kinase kinase 2 regulate androgen receptor activity in castrate resistant prostate cancer cells.

Re-activation of the transcriptional activity of the androgen receptor (AR) is an important factor mediating progression from androgen-responsive to castrate-resistant prostate cancer (CRPC). However, the mechanisms regulating AR activity in CRPC remain incompletely understood. Ca(2+) /calmodulin-dependent kinase kinase (CaMKK) 2 was previously shown to regulate AR activity in androgen-responsive prostate cancer cells. Our objective was to further explore the basis of this regulation in CRPC cells. The abundance of CaMKK2 in nuclear fractions of androgen-responsive prostate cancer and CRPC, cells were determined by subcellular fractionation and Western blotting. CaMKK2 association with nuclear pore complexes (NPCs) and nucleoporins (Nups) including Nup62, were imaged by structured illumination and super-resolution fluorescence microscopy and co-immunoprecipitation, respectively. The abundance and subcellular localization of CaMKK2 and Nup62 in human clinical specimens of prostate cancer was visualized by immunohistochemistry. The role of Nups in the growth and viability of CRPC cells was assessed by RNA interference and cell counting. The involvement of CaMKK2 and Nup62 in regulating AR transcriptional activity was addressed by RNA interference, chromatin immunoprecipitation, androgen response element reporter assay, and Western blotting. CaMKK2 was expressed at higher levels in the nuclear fraction of CPRC C4-2 cells, than in that of androgen-responsive LNCaP cells. In C4-2 cells, CaMKK2 associated with NPCs of the nuclear envelope and physically interacted with Nup62. CaMKK2 and Nup62 demonstrated pronounced, and similar increases in both expression and perinuclear/nuclear localization in human clinical specimens of advanced prostate cancer relative to normal prostate. Knockdown of Nup62, but not of Nups, 98 or 88, reduced growth and viability of C4-2 cells. Knockdown of Nup62 produced a greater reduction of the growth and viability of C4-2 cells than of non-neoplastic RWPE-1 prostatic cells. Nup62, CaMKK2, and the AR were recruited to androgen response elements of the AR target genes, prostate specific antigen, and transmembrane protease, serine 2. Knockdown of CaMKK2 and Nup62 reduced prostate specific antigen expression and AR transcriptional activity driven by androgen response elements from the prostate-specific probasin gene promoter. Nup62 and CaMKK2 are required for optimal AR transcriptional activity and a potential mechanism for AR re-activation in CRPC.

Karacosta LG ，Kuroski LA ，Hofmann WA ，Azabdaftari G ，Mastri M ，Gocher AM ，Dai S ，Hoste AJ ，Edelman AM ... -  《-》

A regulatory feedback loop between Ca2+/calmodulin-dependent protein kinase kinase 2 (CaMKK2) and the androgen receptor in prostate cancer progression.

Karacosta LG ，Foster BA ，Azabdaftari G ，Feliciano DM ，Edelman AM ... -  《-》

Regulation of the transcriptional coactivator FHL2 licenses activation of the androgen receptor in castrate-resistant prostate cancer.

McGrath MJ ，Binge LC ，Sriratana A ，Wang H ，Robinson PA ，Pook D ，Fedele CG ，Brown S ，Dyson JM ，Cottle DL ，Cowling BS ，Niranjan B ，Risbridger GP ，Mitchell CA ... -  《-》

Down-regulation of calcium/calmodulin-dependent protein kinase kinase 2 by androgen deprivation induces castration-resistant prostate cancer.

Conversion into androgen-hypersensitive state and adaptation to the low concentration of androgen during ADT cause relapse of prostate cancer (PCa). It is important to identify differentially expressed genes between PCa and normal prostate tissues and to reveal the function of these genes that are involved in progression of PCa. We performed cDNA microarray analysis to identify differentially expressed genes, calcium/calmodulin-dependent protein kinase kinase 2 (CAMKK2). Immunohistochemical analysis was conducted to investigate the relationship between the CAMKK2 expression level and prognosis. The function of CAMKK2 was assessed by generating CAMKK2 overexpressed LNCaP cells and by knockdown of CAMKK2. We identified CAMKK2 overexpressed six times in PCa more than normal prostate by cDNA microarray analysis. Immunohistochemical analysis of CAMKK2 protein showed that CAMKK2 protein was expressed more in PCa than normal tissue. However, the expression in the high-grade PCa diminished. Moreover, the narrowness of CAMKK2-positive area before ADT was a poor prognostic factor. Androgen-deprivation treatment from the medium in which LNCaP cells were cultured in the presence of 10 nM DHT repressed CAMKK2 expression. CAMKK2 overexpressed LNCaP cells (LNCaP/GFP-CAMKK2) attenuated androgen-sensitivity. Tumorigenesis of LNCaP/GFP-CAMKK2 cells in male SCID mice was decreased compared with control cells irrespective of castration. Finally, knockdown of CAMKK2 mRNA in LNCaP cells induced androgen-hypersensitivity and stimulated LNCaP cell proliferation. Induction of androgen-hypersensitivity after ADT may be involved in down-regulation of CAMKK2. This result may provide new therapeutic approach to keep androgen-sensitivity of PCa after ADT.

Shima T ，Mizokami A ，Miyagi T ，Kawai K ，Izumi K ，Kumaki M ，Ofude M ，Zhang J ，Keller ET ，Namiki M ... -  《-》

Lipocalin 2 over-expression facilitates progress of castration-resistant prostate cancer via improving androgen receptor transcriptional activity.

Ding G ，Wang J ，Feng C ，Jiang H ，Xu J ，Ding Q ... -  《Oncotarget》