title: Time-resolved human kinome RNAi screen identifies a network regulating mitotic-events as early regulators of cell proliferation.
abstract: Analysis of biological processes is frequently performed with the help of phenotypic assays where data is mostly acquired in single end-point analysis. Alternative phenotypic profiling techniques are desired where time-series information is essential to the biological question, for instance to differentiate early and late regulators of cell proliferation in loss-of-function studies. So far there is no study addressing this question despite of high unmet interests, mostly due to the limitation of conventional end-point assaying technologies. We present the first human kinome screen with a real-time cell analysis system (RTCA) to capture dynamic RNAi phenotypes, employing time-resolved monitoring of cell proliferation via electrical impedance. RTCA allowed us to investigate the dynamics of phenotypes of cell proliferation instead of using conventional end-point analysis. By introducing data transformation with first-order derivative, i.e. the cell-index growth rate, we demonstrate this system suitable for high-throughput screenings (HTS). The screen validated previously identified inhibitor genes and, additionally, identified activators of cell proliferation. With the information of time kinetics available, we could establish a network of mitotic-event related genes to be among the first displaying inhibiting effects after RNAi knockdown. The time-resolved screen captured kinetics of cell proliferation caused by RNAi targeting human kinome, serving as a resource for researchers. Our work establishes RTCA technology as a novel robust tool with biological and pharmacological relevance amenable for high-throughput screening.
authors.affiliation: Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
authors.forename: Jitao David
authors.initials: JD
authors.lastname: Zhang
authors.forename: Cindy
authors.initials: C
authors.lastname: Koerner
authors.forename: Stephanie
authors.initials: S
authors.lastname: Bechtel
authors.forename: Christian
authors.initials: C
authors.lastname: Bender
authors.forename: Ioanna
authors.initials: I
authors.lastname: Keklikoglou
authors.forename: Christian
authors.initials: C
authors.lastname: Schmidt
authors.forename: Anja
authors.initials: A
authors.lastname: Irsigler
authors.forename: Ute
authors.initials: U
authors.lastname: Ernst
authors.forename: Ozgür
authors.initials: O
authors.lastname: Sahin
authors.forename: Stefan
authors.initials: S
authors.lastname: Wiemann
authors.forename: Ulrich
authors.initials: U
authors.lastname: Tschulena
mesh_terms: D049109:Cell Proliferation / Q000187:drug effects; D057075:Enzyme Assays; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D008938:Mitosis / Q000187:drug effects / Q000235:genetics; D010641:Phenotype; D010770:Phosphotransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D059010:Single-Cell Analysis; D013997:Time Factors
chemical_list: D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; D010770:Phosphotransferases