Functional screening of mammalian mechanosensitive genes using Drosophila RNAi library- Smarcd3/Bap60 is a mechanosensitive pro-inflammatory gene.
Disturbed blood flow (d-flow) induces atherosclerosis by altering the expression of mechanosensitive genes in the arterial endothelium. Previously, we identified >580 mechanosensitive genes in the mouse arterial endothelium, but their role in endothelial inflammation is incompletely understood. From this set, we obtained 84 Drosophila RNAi lines that silences the target gene under the control of upstream activation sequence (UAS) promoter. These lines were crossed with C564-GAL4 flies expressing GFP under the control of drosomycin promoter, an NF-κB target gene and a marker of pathogen-induced inflammation. Silencing of psmd12 or ERN1 decreased infection-induced drosomycin expression, while Bap60 silencing significantly increased the drosomycin expression. Interestingly, knockdown of Bap60 in adult flies using temperature-inducible Bap60 RNAi (C564ts-GAL4-Bap60-RNAi) enhanced drosomycin expression upon Gram-positive bacterial challenge but the basal drosomycin expression remained unchanged compared to the control. In the mammalian system, smarcd3 (mammalian ortholog of Bap60) expression was reduced in the human- and mouse aortic endothelial cells exposed to oscillatory shear in vitro as well as in the d-flow regions of mouse arterial endothelium in vivo. Moreover, siRNA-mediated knockdown of smarcd3 induced endothelial inflammation. In summary, we developed an in vivo Drosophila RNAi screening method to identify flow-sensitive genes that regulate endothelial inflammation.
Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA, USA.
Sandeep
S
Kumar
Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA, USA.
In-Hwan
IH
Jang
Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA, USA.
Chan Woo
CW
Kim
Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA, USA.
Dong-Won
DW
Kang
National Creative Research Initiative Center for Hologenomics, and School of Biological Sciences, Seoul National University, Seoul 151-742, South Korea.
Won Jae
WJ
Lee
Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, GA, USA.|Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA.
Hanjoong
H
Jo
D000818:Animals; D001011:Aorta / Q000166:cytology; D002478:Cells, Cultured; D002868:Chromosomal Proteins, Non-Histone / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004330:Drosophila / Q000378:metabolism; D029721:Drosophila Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism; D015723:Gene Library; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D007249:Inflammation / Q000209:etiology; D016209:Interleukin-8 / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009124:Muscle Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D033081:Shear Strength; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism
C462944:Bap60 protein, Drosophila; D002868:Chromosomal Proteins, Non-Histone; D029721:Drosophila Proteins; D016209:Interleukin-8; D009124:Muscle Proteins; C494055:Smarcd3 protein, mouse; D014157:Transcription Factors; D019010:Vascular Cell Adhesion Molecule-1; C091227:DRS protein, Drosophila