An endoribonuclease-prepared siRNA screen in human cells identifies genes essential for cell division

Ralf Kittler, Gabriele Putz, Laurence Pelletier, Ina Poser, Anne-Kristin Heninger, David Drechsel, Steffi Fischer, Irena Konstantinova, Bianca Habermann, Hannes Grabner, Marie-Laure Yaspo, Heinz Himmelbauer, Bernd Korn, Karla Neugebauer, Maria Teresa Pisabarro and Frank Buchholz ()
Additional contact information
Ralf Kittler: Max Planck Institute for Molecular Cell Biology and Genetics
Gabriele Putz: Max Planck Institute for Molecular Cell Biology and Genetics
Laurence Pelletier: Max Planck Institute for Molecular Cell Biology and Genetics
Ina Poser: Max Planck Institute for Molecular Cell Biology and Genetics
Anne-Kristin Heninger: Max Planck Institute for Molecular Cell Biology and Genetics
David Drechsel: Max Planck Institute for Molecular Cell Biology and Genetics
Steffi Fischer: Max Planck Institute for Molecular Cell Biology and Genetics
Irena Konstantinova: Max Planck Institute for Molecular Cell Biology and Genetics
Bianca Habermann: Scionics Computer Innovation, GmbH
Hannes Grabner: Max Planck Institute for Molecular Cell Biology and Genetics
Marie-Laure Yaspo: Max Planck Institute for Molecular Genetics
Heinz Himmelbauer: Max Planck Institute for Molecular Genetics
Bernd Korn: RZPD-Ressourcenzentrum für Genomforschung
Karla Neugebauer: Max Planck Institute for Molecular Cell Biology and Genetics
Maria Teresa Pisabarro: Max Planck Institute for Molecular Cell Biology and Genetics
Frank Buchholz: Max Planck Institute for Molecular Cell Biology and Genetics

Nature, 2004, vol. 432, issue 7020, 1036-1040

Abstract: Abstract RNA interference (RNAi) is an evolutionarily conserved defence mechanism whereby genes are specifically silenced through degradation of messenger RNAs; this process is mediated by homologous double-stranded (ds)RNA molecules1,2,3,4. In invertebrates, long dsRNAs have been used for genome-wide screens and have provided insights into gene functions5,6,7,8. Because long dsRNA triggers a nonspecific interferon response in many vertebrates, short interfering (si)RNA or short hairpin (sh)RNAs must be used for these organisms to ensure specific gene silencing9,10,11. Here we report the generation of a genome-scale library of endoribonuclease-prepared short interfering (esi)RNAs12 from a sequence-verified complementary DNA collection representing 15,497 human genes. We used 5,305 esiRNAs from this library to screen for genes required for cell division in HeLa cells. Using a primary high-throughput cell viability screen followed by a secondary high content videomicroscopy assay, we identified 37 genes required for cell division. These include several splicing factors for which knockdown generates mitotic spindle defects. In addition, a putative nuclear-export terminator was found to speed up cell proliferation and mitotic progression after knockdown. Thus, our study uncovers new aspects of cell division and establishes esiRNA as a versatile approach for genomic RNAi screens in mammalian cells.

Date: 2004
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DOI: 10.1038/nature03159

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