title: Fluorescent peptide-PNA chimeras for imaging monoamine oxidase A mRNA in neuronal cells.
abstract: Monoamine oxidases (MAO) catalyze the oxidative deamination of many biogenic amines and are integral proteins found in the mitochondrial outer membrane. Changes in MAO-A levels are associated with depression, trait aggression, and addiction. Here we report the synthesis, characterization, and in vitro evaluation of novel fluorescent peptide-peptide nucleic acid (PNA) chimeras for MAOA mRNA imaging in live neuronal cells. The probes were designed to include MAOA-specific PNA dodecamers, separated by an N-terminal spacer to a μ-opioid receptor targeting peptide (DAMGO), with a spacer and a fluorophore on the C-terminus. The probe was successfully delivered into human SH-SY5Y neuroblastoma cells through μ-opioid receptor-mediated endocytosis. The K(d) by flow cytometry was 11.6 ± 0.8 nM. Uptake studies by fluorescence microscopy showed ∼5-fold higher signal in human SH-SY5Y neuroblastoma cells than in negative control CHO-K1 cells that lack μ-opioid receptors. Moreover, a peptide-mismatch control sequence showed no significant uptake in SH-SY5Y cells. Such mRNA imaging agents with near-infrared fluorophores might enable real time imaging and quantitation of neuronal mRNAs in live animal models.
authors.affiliation: Department of Biochemistry & Molecular Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors.forename: Dalip
authors.initials: D
authors.lastname: Sethi
authors.forename: Chang-Po
authors.initials: CP
authors.lastname: Chen
authors.forename: Rui-Yan
authors.initials: RY
authors.lastname: Jing
authors.forename: Mathew L
authors.initials: ML
authors.lastname: Thakur
authors.forename: Eric
authors.initials: E
authors.lastname: Wickstrom
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000032:analysis* / Q000737:chemistry; D006801:Humans; D008856:Microscopy, Fluorescence; D057054:Molecular Imaging; D015394:Molecular Structure; D008995:Monoamine Oxidase / Q000032:analysis / Q000235:genetics*; D009447:Neuroblastoma / Q000201:enzymology / Q000235:genetics; D009474:Neurons / Q000378:metabolism*; D020135:Peptide Nucleic Acids / Q000032:analysis* / Q000737:chemistry; D010455:Peptides / Q000032:analysis* / Q000737:chemistry; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics / Q000378:metabolism
chemical_list: D005456:Fluorescent Dyes; D020135:Peptide Nucleic Acids; D010455:Peptides; D012333:RNA, Messenger; D008995:Monoamine Oxidase