The mechanisms of INK4-ARF inactivation

Inactivation of the INK4-ARF locus is one of the most frequent events in cancers. The mechanistic basis of epigenetic silencing was not well understood. Recently Gamell et al. found a novel mechanism, in which loss of an E3 ubiquitin ligase E6AP results in the silencing of INK4-ARF locus through E2F-CDC6 pathway.

Cancer is the disease caused by mutations to genes which results in gain of function of oncogene and loss of function of tumor suppressor gene, allowing cells to multiply in an out of control manner. Three well-known tumor suppressors: p14^ARF, p16^INK4A (also known as CDKN2A), and p15^INK4B (also known as CDKN2B) are located in close proximity to one another (within a 35 kb region) at the INK4-ARF locus, yet each is transcribed from a distinct promoter. Interestingly, p14^ARF and p16^INK4A share exons two and three, but each is translated in a different reading frame, yielding unrelated polypeptides. Inactivation of the INK4-ARF locus is one of the most frequent events in cancers (1). It is well established that in many cancers specific genes affecting cellular growth control are hypermethylated and transcriptionally silenced (2,3). Besides of the somatic mutation, the loss of expression of INK4-ARF due to epigenetic alteration often contributes to the initiation and progression of cancers. However, the mechanistic basis of epigenetic silencing was not understood. Through an RNA interference (RNAi) screen, Serra, Fang et al. have identified an oncogene directed transcriptional repression pathway for the silencing of INK4-ARF locus in colorectal cancer (4). The pathway is initiated on DNA by binding of the transcriptional repressor, ZNF304, which recruits a corepressor complex that includes SETDB1, KAP1 and DNMT1, leading to promoter hypermethylation and transcriptional silencing. Activated KRAS regulates the pathway by maintaining high levels of ZNF304, which drives DNA binding.

In a newly published study, an alternative mechanism has been found for the silencing of the INK4-ARF locus involving the E3 ubiquitin ligase and transcriptional cofactor E6AP (also known as UBE3A) (5). In a healthy lung tissue, E6AP induced the expression of the INK4-ARF locus at the transcriptional level by inhibiting CDC6 transcription, a gene encoding a key repressor of the locus in non-small cell lung cancer (NSCLC). In cancer cells, low or lost expression of E6AP results in high abundance of CDC6, which represses the INK4-ARF locus and therefore results in low amounts of p16INK4a, which is tumorigenic in the lung. Loss of p16INK4a or increased CDC6 expression in NSCLC patients is linked to poor prognosis of NSCLC patients. Gamell et al. indicate novel opportunities to therapeutically restore tumor suppression by p16INK4a in NSCLC with the E6AP-low/CDC6-high/p16INK4a-low expression profile.

In summary, the inactivation of INK4-ARF locus is explained in three mechanisms (Figure 1): genetic deletion due to its localization in a common chromosome fragile site FRA9G (6), promoter hypermethylation and transcription silencing by KRAS-ZNF304-DNMT1 pathway (4), transcriptional silencing through low E6AP-high CDC6 axis in a methylation free manner (5).

Figure 1 Three mechanisms for the inactivation of INK4-ARF locus. (A) The frequent genetic deletion of INK4-ARF genes; (B) activated oncogene KRAS stabilizes ZNF304, which drives binding of DNMT1 and its corepressor for the hypermethylation of promoter and silencing of INK4-ARF; (C) loss of E6AP leads to accumulation of repressor CDC6, which inhibits the transcription of INK4-ARF without promoter methylation.

Acknowledgements

None.

Footnote

Conflicts of Interest: The author has no conflicts of interest to declare.

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