title: A High-Throughput Screening Platform Targeting PDLIM5 for Pulmonary Hypertension.
abstract: Pulmonary arterial hypertension is a complex disease with multiple etiologic factors. PDLIM5, a member of the Enigma subfamily of PDZ and LIM domain protein family, contains an N-terminal PDZ domain and three LIM domains at its C-terminus. We have previously shown that overexpression of PDLIM5 prevents hypoxia-induced pulmonary hypertension (PH), and deletion of PDLIM5 in smooth muscle cells enhances hypoxia-induced PH in vivo. These results suggest that PDLIM5 may be a novel therapeutic target of PH. In this study, we aim to establish a high-throughput screening platform for PDLIM5-targeted drug discovery. We generated a stable mink lung epithelial cell line (MLEC) containing a transforming growth factor-β/Smad luciferase reporter with lentivirus-mediated suppression of PDLIM5 (MLEC-shPDLIM5) and measured levels of Smad2/3 and pSmad2/3. We found that in MLEC, suppression of PDLIM5 decreased Smad-dependent luciferase activity, Smad3, and pSmad3. We used MLEC-shPDLIM5 and a control cell line (MLEC-shCTL) to screen the Prestwick library (1200 compounds) and identified and validated paclitaxel as a PDLIM5 inhibitor in MLEC. Furthermore, we showed that paclitaxel inhibited Smad2 expression and Smad3 phosphorylation in A549 cells. Our study suggests that this system is robust and suitable for PDLIM5-targeted drug discovery.
authors.affiliation: Department of Microbiology and Immunology, College of Medicine University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Han
authors.initials: H
authors.lastname: Cheng
authors.affiliation: Department of Pediatrics, College of Medicine University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Tianji
authors.initials: T
authors.lastname: Chen
authors.affiliation: Department of Pediatrics, College of Medicine University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Merve
authors.initials: M
authors.lastname: Tor
authors.affiliation: Department of Pediatrics, College of Medicine University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Deborah
authors.initials: D
authors.lastname: Park
authors.affiliation: Department of Pediatrics, College of Medicine University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Qiyuan
authors.initials: Q
authors.lastname: Zhou
authors.affiliation: Department of Pediatrics, College of Medicine University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Jason B
authors.initials: JB
authors.lastname: Huang
authors.affiliation: Department of Pediatrics, College of Medicine University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Nour
authors.initials: N
authors.lastname: Khatib
authors.affiliation: Department of Microbiology and Immunology, College of Medicine University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Lijun
authors.initials: L
authors.lastname: Rong
authors.affiliation: Department of Pediatrics, College of Medicine University of Illinois at Chicago, Chicago, IL, USA University of Illinois Cancer Center, Chicago, IL, USA guofei@uic.edu.
authors.forename: Guofei
authors.initials: G
authors.lastname: Zhou
mesh_terms: D000072283:A549 Cells; D048868:Adaptor Proteins, Signal Transducing / Q000819:agonists* / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D000818:Animals; D000959:Antihypertensive Agents / Q000494:pharmacology*; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D015870:Gene Expression; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D057166:High-Throughput Screening Assays; D006801:Humans; D006976:Hypertension, Pulmonary / Q000188:drug therapy; D060588:LIM Domain Proteins / Q000819:agonists* / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D016086:Lentivirus / Q000235:genetics / Q000378:metabolism; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008168:Lung / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D008907:Mink; D017239:Paclitaxel / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D051899:Smad2 Protein / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D051900:Smad3 Protein / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000959:Antihypertensive Agents; D060588:LIM Domain Proteins; C512485:PDLIM5 protein, human; D034741:RNA, Small Interfering; C497782:SMAD2 protein, human; C497790:SMAD3 protein, human; D051899:Smad2 Protein; D051900:Smad3 Protein; D016212:Transforming Growth Factor beta; D008156:Luciferases; D017239:Paclitaxel
keywords: PDLIM5; TGF/Smad; high-throughput screening; luciferase reporter assay