title: MiRNA mimic screen for improved expression of functional neurotensin receptor from HEK293 cells.
abstract: Obtaining adequate quantities of functional mammalian membrane proteins has been a bottleneck in their structural and functional studies because the expression of these proteins from mammalian cells is relatively low. To explore the possibility of enhancing expression of these proteins using miRNA, a stable T-REx-293 cell line expressing the neurotensin receptor type 1 (NTSR1), a hard-to-express G protein-coupled receptor (GPCR), was constructed. The cell line was then subjected to human miRNA mimic library screening. In parallel, an HEK293 cell line expressing luciferase was also screened with the same human miRNA mimic library. Five microRNA mimics: hsa-miR-22-5p, hsa-miR-18a-5p, hsa-miR-22-3p, hsa-miR-429, and hsa-miR-2110were identified from both screens. They led to 48% increase in the expression of functional NTSR1 and to 239% increase of luciferase expression. These miRNAs were also effective in enhancing the expression of secretedglypican-3 hFc-fusion protein from HEK293 cells.The results indicate that these molecules may have a wide role in enhancing the production of proteins with biomedical interest.
authors: Biotechnology Core Laboratory, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, 20892.|Departments of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, Maryland.
authors: Su
authors: S
authors: Xiao
authors: NIH Chemical Genomics Center, National Center for Advancing Translational Sciences, National Institutes of health, Rockville, Maryland, 20850.
authors: Yu-Chi
authors: YC
authors: Chen
authors: Departments of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, Maryland.
authors: Michael J
authors: MJ
authors: Betenbaugh
authors: NIH Chemical Genomics Center, National Center for Advancing Translational Sciences, National Institutes of health, Rockville, Maryland, 20850. martinsc@mail.nih.gov.
authors: Scott E
authors: SE
authors: Martin
authors: Biotechnology Core Laboratory, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, 20892. josephs@intra.niddk.nih.gov.
authors: Joseph
authors: J
authors: Shiloach
mesh_terms: D015870:Gene Expression; D057809:HEK293 Cells / Q000502:physiology*; D006801:Humans; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D008403:Mass Screening; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D018028:Receptors, Neurotensin / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D035683:MicroRNAs; D018028:Receptors, Neurotensin; D008156:Luciferases
keywords: HEK293; microRNA screen; neurotensin receptor; protein expression