title: A global approach combining proteome analysis and phenotypic screening with RNA interference yields novel apoptosis regulators.
abstract: Global approaches like proteome or transcriptome analyses have been performed extensively to identify candidate genes or proteins involved in biological and pathological processes. Here we describe the identification of proteins implicated in the regulation of apoptosis using proteome analysis and the functional validation of targets by RNA interference. A high-throughput platform for the validation of synthetic small interfering RNAs (siRNAs) by quantitative real-time PCR was established. Genes of the identified factors were silenced by automated siRNA transfection, and their role in apoptotic signaling was investigated. Using this strategy, nine new modulators of apoptosis were identified. A subsequent detailed study demonstrated that hepatoma-derived growth factor (HDGF) is required for TNFalpha-induced release of pro-apoptotic factors from mitochondria. The strategy described here may be used for hypothesis-free, global gene function analysis.
authors.affiliation: RNAx GmbH, Breddiner Weg 5, 13591 Berlin, Germany.
authors.forename: Nikolaus
authors.initials: N
authors.lastname: Machuy
authors.forename: Bernd
authors.initials: B
authors.lastname: Thiede
authors.forename: Krishnaraj
authors.initials: K
authors.lastname: Rajalingam
authors.forename: Christiane
authors.initials: C
authors.lastname: Dimmler
authors.forename: Oliver
authors.initials: O
authors.lastname: Thieck
authors.forename: Thomas F
authors.initials: TF
authors.lastname: Meyer
authors.forename: Thomas
authors.initials: T
authors.lastname: Rudel
mesh_terms: D017209:Apoptosis / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000502:physiology; D019169:Jurkat Cells; D008928:Mitochondria / Q000378:metabolism; D010641:Phenotype; D020543:Proteome / Q000032:analysis* / Q000235:genetics / Q000502:physiology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D036341:Intercellular Signaling Peptides and Proteins; D020543:Proteome; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; C051610:hepatoma-derived growth factor