Post-Translational Regulation of ARF: Perspective in Cancer

Abstract

Tumorigenesis can be induced by various stresses that cause aberrant DNA mutations and unhindered cell proliferation. Under such conditions, normal cells autonomously induce defense mechanisms, thereby stimulating tumor suppressor activation. ARF, encoded by the CDKN2a locus, is one of the most frequently mutated or deleted tumor suppressors in human cancer. The safeguard roles of ARF in tumorigenesis are mainly mediated via the MDM2-p53 axis, which plays a prominent role in tumor suppression. Under normal conditions, low p53 expression is stringently regulated by its target gene, MDM2 E3 ligase, which induces p53 degradation in a ubiquitin-proteasome-dependent manner. Oncogenic signals induced by MYC, RAS, and E2Fs trap MDM2 in the inhibited state by inducing ARF expression as a safeguard measure, thereby activating the tumor-suppressive function of p53. In addition to the MDM2-p53 axis, ARF can also interact with diverse proteins and regulate various cellular functions, such as cellular senescence, apoptosis, and anoikis, in a p53-independent manner. As the evidence indicating ARF as a key tumor suppressor has been accumulated, there is growing evidence that ARF is sophisticatedly fine-tuned by the diverse factors through transcriptional and post-translational regulatory mechanisms. In this review, we mainly focused on how cancer cells employ transcriptional and post-translational regulatory mechanisms to manipulate ARF activities to circumvent the tumor-suppressive function of ARF. We further discussed the clinical implications of ARF in human cancer.

Keywords: ARF; cancer; p14; phosphorylation; post-translational modification; transcriptional regulation; tumor suppressor; ubiquitination.

Figure 1

Overview of the genomic structure and function of the CDKN2a locus. (A) Two transcripts, α-transcript (encoding INK4a) and β-transcript (encoding ARF), are transcribed from the CDKN2a locus in response to oncogenic stresses. Although these two transcripts share exon 2 and 3 sequences, they have alternative reading frames, and thus are translated into two different proteins. (B) INK4a inhibits cyclin-dependent kinase 4/6 (CDK4/CDK6) activity, leading to an increase in hypo-phosphorylated retinoblastoma (RB) levels. Hypo-phosphorylated RB blocks E2F function, subsequently inducing cell cycle arrest. ARF binds to mouse double minute 2 homolog (MDM2), which accumulates in the nucleolus and inhibits E3 ligase activity. This leads to p53 stabilization, inducing cell cycle arrest and apoptosis. ARF induces apoptosis, cell cycle arrest, and senescence in a p53-independent manner.

Figure 2

A number of transcriptional factors positively or negatively regulate ARF transcription. Smads, DMP1α, E2Fs, MYC, and FoxO activate ARF transcription. E2F3b, enhancer of zeste homolog 2 (Ezh2)/Twist-1, chromobox protein homolog 7 (CBX7), T-box transcription factor 2 (TBX2), B-cell-specific Moloney murine leukemia virus integration site 1 (BMI-1), and epidermal growth factor receptor (EGFR) directly binds to the ARF promoter and suppress ARF transcription. DMP1β interacts with DMP1α, subsequently blocking the binding of DMP1α to the ARF promoter. Furthermore, the transforming growth factor beta 1 (TGFβ1) signaling pathway negatively regulates ARF transcription by inhibiting E2F1 expression.

Figure 3

ARF function, stability, and localization are regulated by phosphorylation, ubiquitination, PPIs, and chaperone-mediated autophagy (CMA). PKCα-induced ARF phosphorylation leads to the cytoplasmic localization of ARF, thereby promoting cell spreading and alleviating anoikis. Makorin ring finger protein 1 (MKRN1), Siva1, and ubiquitin ligase for ARF 1 (ULF1) ubiquitinate ARF, leading to its proteasome-mediated degradation. Ubiquitin-specific peptidase 10 (USP10) directly binds to ARF and then detaches ubiquitin from ARF, thus stabilizing ARF. Proapoptotic nuclear protein 1 (PANO), nucleophosmin (NPM), and tat-binding protein-1 (TBP-1) can bind to ARF, thereby preventing its degradation. Proteasome activator complex subunit 3 (REG-γ) and MDM2 interact with ARF and then transport ARF to the proteasome, thereby resulting in ARF degradation. C-terminus heat shock cognate 71 kDa protein (HSC70)-interacting protein (CHIP)-heat shock protein 90 (HSP90) forms a complex with ARF, leading to degradation of ARF in a lysosome-dependent manner.