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miR-125b targets erythropoietin and its receptor and their expression correlates with metastatic potential and
Manuela Ferracin1, Cristian Bassi, Massimo Pedriali
1Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy. manuela.ferracin@unife.it.
Background:
The microRNA 125b is a double-faced gene expression regulator described both as a tumor suppressor gene (in solid tumors) and an oncogene (in hematologic malignancies). In human breast cancer, it is one of the most down-regulated miRNAs and is able to modulate ERBB2/3 expression. Here, we investigated its targets in breast cancer cell lines after miRNA-mimic transfection. We examined the interactions of the validated targets with ERBB2 oncogene and the correlation of miR-125b expression with clinical variables.
Methods:
MiR-125b possible targets were identified after transfecting a miRNA-mimic in MCF7 cell line and analyzing gene expression modifications with Agilent microarrays and Sylamer bioinformatic tool. Erythropoietin (EPO) and its receptor (EPOR) were validated as targets of miR-125b by luciferase assay and their expression was assessed by RT-qPCR in 42 breast cancers and 13 normal samples. The molecular talk between EPOR and ERBB2 transcripts, through miR-125b, was explored transfecting MDA-MD-453 and MDA-MB-157 with ERBB2 RNA and using RT-qPCR.
Results:
We identified a panel of genes down-regulated after miR-125b transfection and putative targets of miR-125b. Among them, we validated erythropoietin (EPO) and its receptor (EPOR) - frequently overexpressed in breast cancer--as true targets of miR-125b. Moreover, we explored possible correlations with clinical variables and we found a down-regulation of miR-125b in metastatic breast cancers and a significant positive correlation between EPOR and ERBB2/HER2 levels, that are both targets of miR-125b and function as competing endogenous RNAs (ceRNAs).
Conclusions:
Taken together our results show a mechanism for EPO/EPOR and ERBB2 co-regulation in breast cancer and confirm the importance of miR-125b in controlling clinically-relevant cancer features.
Insights
MicroRNA 125b (miR-125b) regulates erythropoietin (EPO) and its receptor (EPOR) in breast cancer. Down-regulation of miR-125b correlates with metastasis and ERBB2/HER2 co-regulation.
Area of Science:
- Molecular Biology
- Genetics
- Oncology
Background:
- MicroRNA 125b (miR-125b) exhibits dual roles as a tumor suppressor or oncogene depending on cancer type.
- In human breast cancer, miR-125b is frequently downregulated and influences ERBB2/3 expression.
Purpose of the Study:
- To identify miR-125b targets in breast cancer cell lines.
- To investigate the interaction between miR-125b targets, ERBB2, and clinical variables.
Main Methods:
- MiRNA mimic transfection in MCF7 cells followed by gene expression analysis (Agilent microarrays, Sylamer).
- Luciferase assays and RT-qPCR to validate erythropoietin (EPO) and EPOR as miR-125b targets.
- ERBB2 RNA transfection and RT-qPCR to study EPOR and ERBB2 transcript interactions.
Main Results:
- Identified several genes downregulated by miR-125b, validating EPO and EPOR as direct targets.
- Found miR-125b downregulation in metastatic breast cancers.
- Observed a positive correlation between EPOR and ERBB2/HER2 levels, identifying them as competing endogenous RNAs (ceRNAs).
Conclusions:
- Established a co-regulation mechanism for EPO/EPOR and ERBB2 in breast cancer via miR-125b.
- Confirmed miR-125b's critical role in regulating key cancer features and potential therapeutic implications.
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