title: 14-3-3 isoforms bind directly exon B of the 5'-UTR of human surfactant protein A2 mRNA.
abstract: Human surfactant protein (SP) A (SP-A), an innate immunity molecule, is encoded by two genes, SFTPA1 and SFTPA2. The 5'-untranslated splice variant of SP-A2 (ABD), but not SP-A1 (AD), contains exon B (eB). eB is an enhancer for transcription and translation and contains cis-regulatory elements. Specific trans-acting factors, including 14-3-3, bind eB. The 14-3-3 protein family contains seven isoforms that have been found by mass spectrometry in eB electromobility shift assays (Noutsios et al. Am J Physiol Lung Cell Mol Physiol 304: L722-L735, 2013). We used four different approaches to investigate whether 14-3-3 isoforms bind directly to eB. 1) eB RNA pulldown assays showed that 14-3-3 isoforms specifically bind eB. 2) RNA electromobility shift assay complexes were formed using purified 14-3-3 isoforms β, γ, ε, η, σ, and τ, but not isoform ζ, with wild-type eB RNA. 3 and 4) RNA affinity chromatography assays and surface plasmon resonance analysis showed that 14-3-3 isoforms β, γ, ε, η, σ, and τ, but not isoform ζ, specifically and directly bind eB. Inhibition of 14-3-3 isoforms γ, ε, η, and τ/θ with shRNAs in NCI-H441 cells resulted in downregulation of SP-A2 levels but did not affect SP-A1 levels. However, inhibition of 14-3-3 isoform σ was correlated with lower levels of SP-A1 and SP-A2. Inhibition of 14-3-3 isoform ζ/δ, which does not bind eB, had no effect on expression levels of SP-A1 and SP-A2. In conclusion, the 14-3-3 protein family affects differential regulation of SP-A1 and SP-A2 by binding directly to SP-A2 5'-UTR mRNA.
authors.affiliation: Center for Host Defense, Inflammation, and Lung Disease (CHILD), Research Department of Pediatrics, College of Medicine, The Pennsylvania State University, Hershey, Pennsylvania; and.
authors.forename: Georgios T
authors.initials: GT
authors.lastname: Noutsios
authors.affiliation: Center for Host Defense, Inflammation, and Lung Disease (CHILD), Research Department of Pediatrics, College of Medicine, The Pennsylvania State University, Hershey, Pennsylvania; and.
authors.forename: Paul
authors.initials: P
authors.lastname: Ghattas
authors.affiliation: Center for Host Defense, Inflammation, and Lung Disease (CHILD), Research Department of Pediatrics, College of Medicine, The Pennsylvania State University, Hershey, Pennsylvania; and.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Bennett
authors.affiliation: Center for Host Defense, Inflammation, and Lung Disease (CHILD), Research Department of Pediatrics, College of Medicine, The Pennsylvania State University, Hershey, Pennsylvania; and Department of Obstetrics and Gynecology, College of Medicine, The Pennsylvania State University, Hershey, Pennsylvania jfloros@psu.edu.
authors.forename: Joanna
authors.initials: J
authors.lastname: Floros
mesh_terms: D048948:14-3-3 Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020121:5' Untranslated Regions / Q000235:genetics*; D000230:Adenocarcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D005091:Exons / Q000235:genetics*; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D020033:Protein Isoforms; D037662:Pulmonary Surfactant-Associated Protein A / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
chemical_list: D048948:14-3-3 Proteins; D020121:5' Untranslated Regions; D020033:Protein Isoforms; D037662:Pulmonary Surfactant-Associated Protein A; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C445843:SFTPA2 protein, human
keywords: 14-3-3; 5′-untranslated region; human pulmonary surfactant protein A; translation