title: MicroRNA-31 controls G protein alpha-13 (GNA13) expression and cell invasion in breast cancer cells.
abstract: BACKGROUND
  Gα13 (GNA13) is the α subunit of a heterotrimeric G protein that mediates signaling through specific G protein-coupled receptors (GPCRs). Our recent study showed that control of GNA13 expression by specific microRNAs (miRNAs or miRs) is important for prostate cancer cell invasion. However, little is known about the control of GNA13 expression in breast cancers. This project was carried out to determine (i) whether enhanced GNA13 expression is important for breast cancer cell invasion, and (ii) if so, the mechanism of deregulation of GNA13 expression in breast cancers.
  
  
  METHODS
  To determine the probable miRNAs regulating GNA13, online miRNA target prediction tool Targetscan and Luciferase assays with GNA13-3'-UTR were used. Effect of miRNAs on GNA13 mRNA, protein and invasion was studied using RT-PCR, western blotting and in vitro Boyden chamber assay respectively. Cell proliferation was done using MTT assays.
  
  
  RESULTS
  Overexpression of GNA13 in MCF-10a cells induced invasion, whereas knockdown of GNA13 expression in MDA-MB-231 cells inhibited invasion. Expression analysis of miRNAs predicted to bind the 3'-UTR of GNA13 revealed that miR-31 exhibited an inverse correlation to GNA13 protein expression in breast cancer cells. Ectopic expression of miR-31 in MDA-MB-231 cells significantly reduced GNA13 mRNA and protein levels, as well as GNA13-3'-UTR-reporter activity. Conversely, blocking miR-31 activity in MCF-10a cells induced GNA13 mRNA, protein and 3'-UTR reporter activity. Further, expression of miR-31 significantly inhibited MDA-MB-231 cell invasion, and this effect was partly rescued by ectopic expression of GNA13 in these cells. Examination of 48 human breast cancer tissues revealed that GNA13 mRNA levels were inversely correlated to miR-31 levels.
  
  
  CONCLUSIONS
  These data provide strong evidence that GNA13 expression in breast cancer cells is regulated by post-transcriptional mechanisms involving miR-31. Additionally our data shows that miR-31 regulates breast cancer cell invasion partially via targeting GNA13 expression in breast cancer cells. Loss of miR-31 expression and increased GNA13 expression could be used as biomarkers of breast cancer progression.
authors.affiliation: Program in Cancer and Stem Cell Biology, Duke-NUS Graduate Medical School, 8 College Road, 169857, Singapore, Singapore. kabeer.rasheed@duke-nus.edu.sg.
authors.forename: Suhail Ahmed Kabeer
authors.initials: SA
authors.lastname: Rasheed
authors.affiliation: Program in Cancer and Stem Cell Biology, Duke-NUS Graduate Medical School, 8 College Road, 169857, Singapore, Singapore. cuirong.teo@duke-nus.edu.sg.
authors.forename: Cui Rong
authors.initials: CR
authors.lastname: Teo
authors.affiliation: Program in Cancer and Stem Cell Biology, Duke-NUS Graduate Medical School, 8 College Road, 169857, Singapore, Singapore. emmanuelbeillard@gmail.com.
authors.forename: Emmanuel Jean
authors.initials: EJ
authors.lastname: Beillard
authors.affiliation: Program in Cancer and Stem Cell Biology, Duke-NUS Graduate Medical School, 8 College Road, 169857, Singapore, Singapore. mathijs.voorhoeve@gmail.com.
authors.forename: P Mathijs
authors.initials: PM
authors.lastname: Voorhoeve
authors.affiliation: Program in Cardiovascular and Metabolic Disorders, Duke-NUS Graduate Medical School, 8 College Road, 169857, Singapore, Singapore. zhou.wei@duke-nus.edu.sg.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhou
authors.affiliation: Centre for Computational Biology & Program in Cardiovascular and Metabolic Disorders, Duke-NUS Graduate Medical School, 169857, Singapore, Singapore. sujoy.ghosh@duke-nus.edu.sg.
authors.forename: Sujoy
authors.initials: S
authors.lastname: Ghosh
authors.affiliation: Program in Cancer and Stem Cell Biology, Duke-NUS Graduate Medical School, 8 College Road, 169857, Singapore, Singapore. patrick.casey@duke-nus.edu.sg.
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Casey
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D051702:Activating Transcription Factor 6; D050976:Basic-Leucine Zipper Transcription Factors / Q000235:genetics*; D014408:Biomarkers, Tumor / Q000235:genetics; D001940:Breast / Q000473:pathology; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D049109:Cell Proliferation / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D009361:Neoplasm Invasiveness / Q000235:genetics* / Q000473:pathology; D012323:RNA Processing, Post-Transcriptional / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics
chemical_list: D020413:3' Untranslated Regions; C487807:ATF6B protein, human; D051702:Activating Transcription Factor 6; D050976:Basic-Leucine Zipper Transcription Factors; D014408:Biomarkers, Tumor; C534612:MIRN31 microRNA, human; D035683:MicroRNAs; D012333:RNA, Messenger