title: Cell-based phenotypic screening of mast cell degranulation unveils kinetic perturbations of agents targeting phosphorylation.
abstract: Mast cells play an essential role in initiating allergic diseases. The activation of mast cells are controlled by a complicated signal network of reversible phosphorylation, and finding the key regulators involved in this network has been the focus of the pharmaceutical industry. In this work, we used a method named Time-dependent cell responding profile (TCRP) to track the process of mast cell degranulation under various perturbations caused by agents targeting phosphorylation. To test the feasibility of this high-throughput cell-based phenotypic screening method, a variety of biological techniques were used. We further screened 145 inhibitors and clustered them based on the similarities of their TCRPs. Stat3 phosphorylation has been widely reported as a key step in mast cell degranulation. Interestingly, our TCRP results showed that a Stat3 inhibitor JSI124 did not inhibit degranulation like other Stat3 inhibitors, such as Stattic, clearly inhibited degranulation. Regular endpoint assays demonstrated that the distinctive TCRP of JSI124 potentially correlated with the ability to induce apoptosis. Consequently, different agents possibly have disparate functions, which can be conveniently detected by TCRP. From this perspective, our TCRP screening method is reliable and sensitive when it comes to discovering and selecting novel compounds for new drug developments.
authors: Program in Molecular Cell Biology, Department of Basic Medical Sciences, Zhejiang University School of Medicine, Hangzhou 310058, China.
authors: Shenlu
authors: S
authors: Qin
authors: Program in Molecular Cell Biology, Department of Basic Medical Sciences, Zhejiang University School of Medicine, Hangzhou 310058, China.
authors: Xumeng
authors: X
authors: Wang
authors: Program in Molecular Cell Biology, Department of Basic Medical Sciences, Zhejiang University School of Medicine, Hangzhou 310058, China.
authors: Huanwen
authors: H
authors: Wu
authors: Program in Molecular Cell Biology, Department of Basic Medical Sciences, Zhejiang University School of Medicine, Hangzhou 310058, China.
authors: Peng
authors: P
authors: Xiao
authors: Program in Molecular Cell Biology, Department of Basic Medical Sciences, Zhejiang University School of Medicine, Hangzhou 310058, China.
authors: Hongqiang
authors: H
authors: Cheng
authors: Program in Molecular Cell Biology, Department of Basic Medical Sciences, Zhejiang University School of Medicine, Hangzhou 310058, China.
authors: Xue
authors: X
authors: Zhang
authors: Program in Molecular Cell Biology, Department of Basic Medical Sciences, Zhejiang University School of Medicine, Hangzhou 310058, China.
authors: Yuehai
authors: Y
authors: Ke
mesh_terms: D000818:Animals; D017209:Apoptosis; D015550:Cell Degranulation; D045744:Cell Line, Tumor; D004247:DNA / Q000737:chemistry; D015195:Drug Design; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004566:Electrodes; D005434:Flow Cytometry; D007073:Immunoglobulin E / Q000737:chemistry; D007700:Kinetics; D008407:Mast Cells / Q000166:cytology*; D010641:Phenotype; D010766:Phosphorylation; D051381:Rats; D050796:STAT3 Transcription Factor / Q000037:antagonists & inhibitors / Q000378:metabolism; D014315:Triterpenes / Q000494:pharmacology
chemical_list: D050796:STAT3 Transcription Factor; C494088:Stat3 protein, rat; D014315:Triterpenes; D007073:Immunoglobulin E; D004247:DNA; C038106:cucurbitacin I