title: Engineering an effective Mn-binding MRI reporter protein by subcellular targeting.
abstract: OBJECTIVE
  Manganese (Mn) is an effective contrast agent and biologically active metal, which has been widely used for Mn-enhanced MRI (MEMRI). The purpose of this study was to develop and test a Mn binding protein for use as a genetic reporter for MEMRI.
  
  
  METHODS
  The bacterial Mn-binding protein, MntR was identified as a candidate reporter protein. MntR was engineered for expression in mammalian cells, and targeted to different subcellular organelles, including the Golgi Apparatus where cellular Mn is enriched. Transfected HEK293 cells and B16 melanoma cells were tested in vitro and in vivo, using immunocytochemistry, MR imaging and relaxometry.
  
  
  RESULTS
  Subcellular targeting of MntR to the cytosol, endoplasmic reticulum and Golgi apparatus was verified with immunocytochemistry. After targeting to the Golgi, MntR expression produced robust R1 changes and T1 contrast in cells, in vitro and in vivo. Co-expression with the divalent metal transporter DMT1, a previously described Mn-based reporter, further enhanced contrast in B16 cells in culture, but in the in vivo B16 tumor model tested was not significantly better than MntR alone.
  
  
  CONCLUSIONS
  This second-generation reporter system both expands the capabilities of genetically encoded reporters for imaging with MEMRI and provides important insights into the mechanisms of Mn biology which create endogenous MEMRI contrast.
authors.affiliation: Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York, USA.|Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Benjamin B
authors.initials: BB
authors.lastname: Bartelle
authors.affiliation: Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York, USA.|Koch Institute of Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Miyeko D
authors.initials: MD
authors.lastname: Mana
authors.affiliation: Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York, USA.
authors.forename: Giselle A
authors.initials: GA
authors.lastname: Suero-Abreu
authors.affiliation: Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York, USA.
authors.forename: Joe J
authors.initials: JJ
authors.lastname: Rodriguez
authors.affiliation: Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York, USA.|Departments of Radiology and Pathology, New York University School of Medicine, New York, New York, USA.
authors.forename: Daniel H
authors.initials: DH
authors.lastname: Turnbull
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D003287:Contrast Media / Q000378:metabolism; D017930:Genes, Reporter / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D008279:Magnetic Resonance Imaging / Q000379:methods*; D008345:Manganese / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D015336:Molecular Probe Techniques; D015335:Molecular Probes / Q000235:genetics / Q000493:pharmacokinetics; D009374:Neoplasms, Experimental / Q000378:metabolism* / Q000473:pathology; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D003287:Contrast Media; C433792:MntR protein, bacteria; D015335:Molecular Probes; D012097:Repressor Proteins; D008345:Manganese
keywords: DMT1; Mn-enhanced MRI (MEMRI); MntR; manganese (Mn); molecular imaging