title: Identification of miRNAs that modulate glucocerebrosidase activity in Gaucher disease cells.
abstract: Gaucher disease is an autosomal recessive disorder caused by deficiency of the enzyme glucocerebrosidase. Although it is a monogenic disease, there is vast phenotypic heterogeneity, even among patients with the same genotype. MicroRNAs (miRNAs) are small non-coding RNAs involved in many biological processes and diseases. To determine whether miRNAs can affect glucocerebrosidase activity, we performed a screen of 875 different miRNA mimics. The screen was performed using Gaucher fibroblasts, and glucocerebrosidase activity was used as the initial outcome parameter. We found several miRNAs that either up- or down-regulated glucocerebrosidase activity. In follow-up assays, we confirmed that one specific miRNA (miR-127-5p) down-regulated both glucocerebrosidase activity and protein levels by down-regulation of LIMP-2, the receptor involved in proper trafficking of glucocerebrosidase from the endoplasmic reticulum to the lysosome. A conditioned media assay demonstrated that cells treated with this miRNA secreted glucocerebrosidase into the extracellular environment, supporting impaired LIMP-2 function. Two other miRNAs, miR-16-5p and miR-195-5p, were found to up-regulate glucocerebrosidase activity by greater than 40% and to enhance expression and protein levels of the enzyme. In conclusion, we show that miRNAs can alter glucocerebrosidase activity in patient cells, indicating that miRNAs can potentially act as modifiers in Gaucher disease.
authors.affiliation: a Section on Molecular Neurogenetics; Medical Genetics Branch; National Human Genome Research Institute; National Institutes of Health ; Bethesda , MD USA.
authors.forename: Marina
authors.initials: M
authors.lastname: Siebert
authors.forename: Wendy
authors.initials: W
authors.lastname: Westbroek
authors.forename: Yu-Chi
authors.initials: YC
authors.lastname: Chen
authors.forename: Nima
authors.initials: N
authors.lastname: Moaven
authors.forename: Yan
authors.initials: Y
authors.lastname: Li
authors.forename: Arash
authors.initials: A
authors.lastname: Velayati
authors.forename: Maria Luiza
authors.initials: ML
authors.lastname: Saraiva-Pereira
authors.forename: Scott E
authors.initials: SE
authors.lastname: Martin
authors.forename: Ellen
authors.initials: E
authors.lastname: Sidransky
mesh_terms: D015153:Blotting, Western; D049109:Cell Proliferation; D002478:Cells, Cultured; D005347:Fibroblasts / Q000201:enzymology*; D005776:Gaucher Disease / Q000201:enzymology / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic; D005962:Glucosylceramidase / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D051907:Lysosomal Membrane Proteins / Q000235:genetics / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D051116:Receptors, Scavenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D051907:Lysosomal Membrane Proteins; D035683:MicroRNAs; D012333:RNA, Messenger; D051116:Receptors, Scavenger; C074299:SCARB2 protein, human; D008156:Luciferases; D005962:Glucosylceramidase
keywords: GBA; GCase; GCase, glucocerebrosidase; GD, Gaucher disease; Gaucher Disease; NegCtrl siRNA, negative control siRNA; UTR, untranslated region; WT, Wild Type; glucocerebrosidase; miRNA; miRNA, microRNA; mimics; screen