AKT-mediated stabilization of histone methyltransferase WHSC1 promotes prostate cancer metastasis
Ni Li, … , Qintong Li, Jun Qin
Ni Li, … , Qintong Li, Jun Qin
Published March 20, 2017
Citation Information: J Clin Invest. 2017;127(4):1284-1302. https://doi.org/10.1172/JCI91144.
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AKT-mediated stabilization of histone methyltransferase WHSC1 promotes prostate cancer metastasis

Abstract

Loss of phosphatase and tensin homolog (PTEN) and activation of the PI3K/AKT signaling pathway are hallmarks of prostate cancer (PCa). However, these alterations alone are insufficient for cells to acquire metastatic traits. Here, we have shown that the histone dimethyl transferase WHSC1 critically drives indolent PTEN-null tumors to become metastatic PCa. In a PTEN-null murine PCa model, WHSC1 overexpression in prostate epithelium cooperated with Pten deletion to produce a metastasis-prone tumor. Conversely, genetic ablation of Whsc1 prevented tumor progression in PTEN-null mice. Molecular characterization revealed that increased AKT activity due to PTEN loss directly phosphorylates WHSC1 at S172, preventing WHSC1 degradation by CRL4Cdt2 E3 ligase. Increased WHSC1 expression transcriptionally upregulates expression of RICTOR, a pivotal component of mTOR complex 2 (mTORC2), to further enhance AKT activity. Therefore, the AKT/WHSC1/mTORC2 signaling cascade represents a vicious feedback loop that elicits unrestrained AKT signaling. Furthermore, we determined that WHSC1 positively regulates Rac1 transcription to increase tumor cell motility. The biological importance of a WHSC1-mediated signaling cascade is substantiated by patient sample analysis in which WHSC1 signaling is tightly correlated with disease progression and recurrence. Taken together, our findings highlight a pivotal link between an epigenetic regulator, WHSC1, and key intracellular signaling molecules, AKT, RICTOR, and Rac1, to drive PCa metastasis.

Authors

Ni Li, Wei Xue, Huairui Yuan, Baijun Dong, Yufeng Ding, Yongfeng Liu, Min Jiang, Shan Kan, Tongyu Sun, Jiale Ren, Qiang Pan, Xiang Li, Peiyuan Zhang, Guohong Hu, Yan Wang, Xiaoming Wang, Qintong Li, Jun Qin

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Figure 5

AKT phosphorylates WHSC1 at the S172 site to stabilize its protein in PCa cells.

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AKT phosphorylates WHSC1 at the S172 site to stabilize its protein in PC...
(A) Western blot analysis of the indicated protein in mouse AP lysates (mAP) and human prostate cell lines (left). qRT-PCR analysis of relative WHSC1 mRNA (right). (B) IB analysis of Ptenfl/fl mouse embryonic fibroblasts (MEF) infected with Ad-GFP or Ad-Cre. Relative Pten and Whsc1 mRNA levels are shown in the right panel. Student’s t test, **P < 0.01. (C) IB analysis of PC3 and LNCaP cells treated with 30 μM LY294002 for the indicated duration of time. (D) IB analysis of PC3 cells transfected with scramble or AKT oligonucleotides with or without MG132 treatment for the indicated duration of time. (E) Sequence alignment of the putative AKT phosphorylation site at S172 in WHSC1. (F) IB analysis of whole cell lysates (WCL) and immunoprecipitates derived from PC3 cells as indicated. (G) In vitro kinase assays depicting major AKT phosphorylation sites in WHSC1. (H) IB analysis of WCL and immunoprecipitates derived from PC3 cells treated with LY294002, IGF-1, or λ-phosphatase. (I) Left: The indicated protein in WT and S172A mutated cells. Right: Relative WHSC1 mRNA expression. Statistical significance was determined by 1-way ANOVA. (J) WT and S172A cell lysates were subjected to IP with anti-WHSC1 antibody and IB with anti-ubiquitin (anti-Ub) and anti-AKT substrate antibody. (K) Representative BLI and x-ray images of the mice after 2 months of inoculations with control and S172A cells (left). Quantitation of BLI and osteolytic area sizes in limbs (right; n = 5). One-way ANOVA followed by Tukey’s multiple comparisons test, **P < 0.01. (L) Representative IHC staining of WHSC1 and phospho-AKT in patients. Box plot shows the relative WHSC1 level in lower, middle, and higher phospho-AKT patients (an Asian radical prostatectomy cohort). Staining indexes using a 10-point quantification scale. Low, <4; middle, 4 and 6; high, >6. Wilcoxon signed-rank test was used to calculate statistical significance. Scale bar: 50 μm (L).