Reduced abundance of the E3 ubiquitin ligase E6AP contributes to decreased expression of the INK4/ARF locus in non–small cell lung cancer

Abstract

The tumor suppressor p16, one protein encoded by the INK4/ARF locus, is frequently absent in multiple cancers, including non-small cell lung cancer (NSCLC). Whereas increased methylation of the encoding gene (CDKN2A) accounts for its loss in a third of patients, no molecular explanation exists for the remainder. We unraveled an alternative mechanism for the silencing of the INK4/ARF locus involving the E3 ubiquitin ligase and transcriptional cofactor E6AP (also known as UBE3A). We found that the expression of three tumor suppressor genes encoded in the INK4/ARF locus (p15, p16, and p19) was decreased in E6AP mouse embryo fibroblasts. 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. Luciferase assays revealed that E6AP inhibited CDC6 expression by reducing its E2F1-dependent transcription. Chromatin immunoprecipitation analysis indicated that E6AP reduced the amount of E2F1 at the CDC6 promoter. In a subset of NSCLC samples, an E6AP-low/CDC6-high/p16-low protein abundance profile correlated with low methylation of the gene encoding p16 (CDKN2A) and poor patient prognosis. These findings define a previously unrecognized tumor-suppressive role for E6AP in NSCLC, reveal an alternative silencing mechanism of the INK4/ARF locus, and reveal E6AP as a potential prognostic marker in NSCLC.

Uncovering a novel pathway for p16 silencing: Therapeutic implications for lung cancer

“…In our recent study published in Science Signaling we addressed, at least in part, this gap in knowledge. 1 We unraveled a novel mechanism for the reduction of p16, involving the E3 ubiquitin ligase and transcriptional cofactor, E6-associated protein (E6AP), which we found to be lowly expressed in certain NSCLCs.…”

“…In our recent study published in Science Signaling we addressed, at least in part, this gap in knowledge. 1 We unraveled a novel mechanism for the reduction of p16, involving the E3 ubiquitin ligase and transcriptional cofactor, E6-associated protein (E6AP), which we found to be lowly expressed in certain NSCLCs.Our analysis identified E6AP as a positive regulator of the INK4/ARF locus. Mechanistically, we showed that E6AP controls this locus by reducing the ability of E2F transcription factor 1 (E2F1) to bind to the cell division control protein 6 (CDC6) promoter.…”

Nano‐LC based proteomic approach identifies that E6AP interacts with ENO1 and targets it for degradation in breast cancer cells

“…Dysregulation of E6AP expression and function has been associated with numerous diseases such as cervical carcinogenesis, Angelman syndrome and others . In a recent study E6AP was shown to act as a tumor suppressor in non‐small cell lung cancer by maintaining INK4/ARF expression levels . Mansour et al showed that E6AP suppressed breast cancer invasiveness, colonization, and metastasis in mice as well as in breast cancer patients while E6AP loss was associated with poor prognosis, particularly for basal breast cancer .…”

Proteotranscriptomic Measurements of E6-Associated Protein (E6AP) Targets in DU145 Prostate Cancer Cells

“…E6AP was initially identified in the context of high risk human papillomavirus (HPV)-related cancers, where E6AP in association with viral oncoprotein E6 ubiquitinates and degrades the tumor suppressor p53 (6). Recent work from our group and others reveal roles of E6AP in HPV-independent cancers, including lung and blood cancer (7,8). E6AP functions as an E3 ligase and as a transcriptional cofactor (9).…”

“…The functions of E6AP, mostly attributed to its ligase activity include cellular growth, cell cycle, apoptosis, cellular senescence, cellular stress response and proteasomal regulation (10 -16). Independent of its E3 ligase activity, E6AP also coactivates transcription by the steroid hormone receptors and by E2F1 (7,9,17).…”