title: miR-125b targets erythropoietin and its receptor and their expression correlates with metastatic potential and ERBB2/HER2 expression.
abstract: 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.
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy. manuela.ferracin@unife.it.
authors.forename: Manuela
authors.initials: M
authors.lastname: Ferracin
authors.forename: Cristian
authors.initials: C
authors.lastname: Bassi
authors.forename: Massimo
authors.initials: M
authors.lastname: Pedriali
authors.forename: Sara
authors.initials: S
authors.lastname: Pagotto
authors.forename: Lucilla
authors.initials: L
authors.lastname: D'Abundo
authors.forename: Barbara
authors.initials: B
authors.lastname: Zagatti
authors.forename: Fabio
authors.initials: F
authors.lastname: Corrà
authors.forename: Gentian
authors.initials: G
authors.lastname: Musa
authors.forename: Elisa
authors.initials: E
authors.lastname: Callegari
authors.forename: Laura
authors.initials: L
authors.lastname: Lupini
authors.forename: Stefano
authors.initials: S
authors.lastname: Volpato
authors.forename: Patrizia
authors.initials: P
authors.lastname: Querzoli
authors.forename: Massimo
authors.initials: M
authors.lastname: Negrini
mesh_terms: D020413:3' Untranslated Regions; D001665:Binding Sites; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D004921:Erythropoietin / Q000235:genetics* / Q000378:metabolism; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D053263:Gene Regulatory Networks; D057809:HEK293 Cells; D006801:Humans; D061986:MCF-7 Cells; D035683:MicroRNAs / Q000235:genetics*; D058977:Molecular Sequence Annotation; D009362:Neoplasm Metastasis; D034622:RNA Interference; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D017467:Receptors, Erythropoietin / Q000235:genetics* / Q000378:metabolism
chemical_list: D020413:3' Untranslated Regions; D004921:Erythropoietin; D035683:MicroRNAs; D018719:Erb-b2 Receptor Tyrosine Kinases; D017467:Receptors, Erythropoietin; C531888:MIRN125 microRNA, human; C508053:ERBB2 protein, human