title: MicroRNA-363 targets myosin 1B to reduce cellular migration in head and neck cancer.
abstract: BACKGROUND
  Squamous cell carcinoma of the head and neck (SCCHN) remains a prevalent and devastating disease. Recently, there has been an increase in SCCHN cases that are associated with high-risk human papillomavirus (HPV) infection. The clinical characteristics of HPV-positive and HPV-negative SCCHN are known to be different but their molecular features are only recently beginning to emerge. MicroRNAs (miRNAs, miRs) are small, non-coding RNAs that are likely to play significant roles in cancer initiation and progression where they may act as oncogenes or tumor suppressors. Previous studies in our laboratory showed that miR-363 is overexpressed in HPV-positive compared to HPV-negative SCCHN cell lines, and the HPV type 16-E6 oncoprotein upregulates miR-363 in SCCHN cell lines. However, the functional role of miR-363 in SCCHN in the context of HPV infection remains to be elucidated.
  
  
  METHODS
  We analyzed miR-363 levels in SCCHN tumors with known HPV-status from The Cancer Genome Atlas (TCGA) and an independent cohort from our institution. Cell migration studies were conducted following the overexpression of miR-363 in HPV-negative cell lines. Bioinformatic tools and a luciferase reporter assay were utilized to confirm that miR-363 targets the 3'-UTR of myosin 1B (MYO1B). MYO1B mRNA and protein expression levels were evaluated following miR-363 overexpression in HPV-negative SCCHN cell lines. Small interfering RNA (siRNA) knockdown of MYO1B was performed to assess the phenotypic implication of reduced MYO1B expression in SCCHN cell lines.
  
  
  RESULTS
  MiR-363 was found to be overexpressed in HPV-16-positive compared to the HPV-negative SCCHN tumors. Luciferase reporter assays performed in HPV-negative JHU028 cells confirmed that miR-363 targets one of its two potential binding sites in the 3'UTR of MYO1B. MYO1B mRNA and protein levels were reduced upon miR-363 overexpression in four HPV-negative SCCHN cell lines. Increased miR-363 expression or siRNA knockdown of MYO1B expression reduced Transwell migration of SCCHN cell lines, indicating that the miR-363-induced migration attenuation of SCCHN cells may act through MYO1B downregulation.
  
  
  CONCLUSIONS
  These findings demonstrate that the overexpression of miR-363 reduces cellular migration in head and neck cancer and reveal the biological relationship between miR-363, myosin 1b, and HPV-positive SCCHN.
authors.affiliation: Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA. bsv4@pitt.edu.|Department of Otolaryngology, University of Pittsburgh and University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA. bsv4@pitt.edu.|Medical Research Fellows Program, Howard Hughes Medical Institute, Chevy Chase, MD, 20815, USA. bsv4@pitt.edu.
authors.forename: Bhavana V
authors.initials: BV
authors.lastname: Chapman
authors.affiliation: Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA. waldai@upmc.edu.
authors.forename: Abigail I
authors.initials: AI
authors.lastname: Wald
authors.affiliation: Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA. paa13@pitt.edu.
authors.forename: Parvez
authors.initials: P
authors.lastname: Akhtar
authors.affiliation: Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA. acv12@pitt.edu.
authors.forename: Ana C
authors.initials: AC
authors.lastname: Munko
authors.affiliation: Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA. Jix58@pitt.edu.
authors.forename: Jingjing
authors.initials: J
authors.lastname: Xu
authors.affiliation: Department of Immunology, University of Pittsburgh, Pittsburgh, PA, 15216, USA. spoveda@pitt.edu.|Department of Otolaryngology, University of Pittsburgh and University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA. spoveda@pitt.edu.
authors.forename: Sandra P
authors.initials: SP
authors.lastname: Gibson
authors.affiliation: Department of Otolaryngology, University of Pittsburgh and University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA. jennifer.grandis@ucsf.edu.|Department of Pharmacology and Chemical Biology, University of Pittsburgh and University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA. jennifer.grandis@ucsf.edu.|Present address: Clinical and Translational Science Institute,, Box 0558, 550 16th Street, 6th Floor, San Francisco, CA, 94158, USA. jennifer.grandis@ucsf.edu.
authors.forename: Jennifer R
authors.initials: JR
authors.lastname: Grandis
authors.affiliation: Department of Immunology, University of Pittsburgh, Pittsburgh, PA, 15216, USA. ferrrl@upmc.edu.|Department of Otolaryngology, University of Pittsburgh and University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA. ferrrl@upmc.edu.
authors.forename: Robert L
authors.initials: RL
authors.lastname: Ferris
authors.affiliation: Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA. khan@pitt.edu.
authors.forename: Saleem A
authors.initials: SA
authors.lastname: Khan
mesh_terms: D020413:3' Untranslated Regions; D000368:Aged; D001483:Base Sequence; D001665:Binding Sites; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D005260:Female; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D006258:Head and Neck Neoplasms / Q000235:genetics* / Q000473:pathology; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000235:genetics*; D008875:Middle Aged; D024461:Myosin Type I / Q000235:genetics*; D009367:Neoplasm Staging; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics
chemical_list: D020413:3' Untranslated Regions; C574861:MIRN363 microRNA, human; C000594736:MYO1B protein, human; D035683:MicroRNAs; D012333:RNA, Messenger; D024461:Myosin Type I