Related Experiment Video
Updated: Mar 13, 2026
Studying Triple Negative Breast Cancer Using Orthotopic Breast Cancer Model
Published on: March 20, 2020
Alternative Polyadenylation in Triple-Negative Breast Tumors Allows NRAS and c-JUN to Bypass PUMILIO
Wayne O Miles1,2, Antonio Lembo3,4, Angela Volorio5
1Department of Molecular Oncology, Massachusetts General Hospital Cancer Center and Harvard Medical School, Charlestown, Massachusetts. dyson@helix.mgh.harvard.edu wayne.miles@osumc.edu.
Abstract:
Alternative polyadenylation (APA) is a process that changes the posttranscriptional regulation and translation potential of mRNAs via addition or deletion of 3' untranslated region (3' UTR) sequences. To identify posttranscriptional-regulatory events affected by APA in breast tumors, tumor datasets were analyzed for recurrent APA events. Motif mapping of the changed 3' UTR regions found that APA-mediated removal of Pumilio regulatory elements (PRE) was unusually common. Breast tumor subtype-specific APA profiling identified triple-negative breast tumors as having the highest levels of APA. To determine the frequency of these events, an independent cohort of triple-negative breast tumors and normal breast tissue was analyzed for APA. APA-mediated shortening of NRAS and c-JUN was seen frequently, and this correlated with changes in the expression of downstream targets. mRNA stability and luciferase assays demonstrated APA-dependent alterations in RNA and protein levels of affected candidate genes. Examination of clinical parameters of these tumors found those with APA of NRAS and c-JUN to be smaller and less proliferative, but more invasive than non-APA tumors. RT-PCR profiling identified elevated levels of polyadenylation factor CSTF3 in tumors with APA. Overexpression of CSTF3 was common in triple-negative breast cancer cell lines, and elevated CSTF3 levels were sufficient to induce APA of NRAS and c-JUN. Our results support the hypothesis that PRE-containing mRNAs are disproportionately affected by APA, primarily due to high sequence similarity in the motifs utilized by polyadenylation machinery and the PUM complex. Cancer Res; 76(24); 7231-41. ©2016 AACR.
Insights
Alternative polyadenylation (APA) shortens mRNA in triple-negative breast tumors, affecting gene expression and tumor invasiveness. This process is linked to elevated CSTF3 levels, driving APA events in cancer.
Area of Science:
- Molecular Biology
- Cancer Research
- Genetics
Background:
- Alternative polyadenylation (APA) regulates mRNA post-transcriptionally by altering 3' untranslated regions (3' UTRs).
- APA impacts mRNA translation and stability, influencing gene expression.
- Recurrent APA events in breast tumors suggest a role in cancer progression.
Purpose of the Study:
- To identify APA-mediated posttranscriptional regulatory events in breast tumors.
- To investigate the association between APA, gene expression, and clinical parameters in breast cancer subtypes.
- To explore the role of polyadenylation factors in driving APA in triple-negative breast cancer.
Main Methods:
- Analysis of breast tumor datasets for recurrent APA events.
- Motif mapping of 3' UTRs to identify regulatory elements affected by APA.
- APA profiling in triple-negative breast tumors and normal breast tissue.
- mRNA stability and luciferase assays to assess APA-dependent gene expression changes.
- RT-PCR and Western blotting to evaluate polyadenylation factor levels.
Main Results:
- APA-mediated removal of Pumilio regulatory elements (PREs) was frequent in breast tumors.
- Triple-negative breast tumors exhibited the highest APA levels.
- APA-mediated shortening of NRAS and c-JUN was observed, correlating with downstream target expression changes.
- Tumors with APA of NRAS and c-JUN showed reduced proliferation but increased invasiveness.
- Elevated CSTF3 levels were associated with APA and sufficient to induce APA of NRAS and c-JUN.
Conclusions:
- APA significantly impacts gene regulation in breast tumors, particularly in triple-negative subtypes.
- APA-driven alterations in NRAS and c-JUN influence tumor characteristics, including invasiveness.
- CSTF3 overexpression is a key driver of APA in triple-negative breast cancer.
- PRE-containing mRNAs are disproportionately affected by APA due to conserved motif similarities.
