title: Activation of RAF1 (c-RAF) by the Marine Alkaloid Lasonolide A Induces Rapid Premature Chromosome Condensation.
abstract: Lasonolide A (LSA), a potent antitumor polyketide from the marine sponge, Forcepia sp., induces rapid and reversible protein hyperphosphorylation and premature chromosome condensation (PCC) at nanomolar concentrations independent of cyclin-dependent kinases. To identify cellular targets of LSA, we screened 2951 shRNAs targeting a pool of human kinases and phosphatases (1140 RefSeqs) to identify genes that modulate PCC in response to LSA. This led to the identification of RAF1 (C-RAF) as a mediator of LSA-induced PCC, as shRNAs against RAF1 conferred resistance to LSA. We found that LSA induced RAF1 phosphorylation on Serine 338 within minutes in human colorectal carcinoma HCT-116, ovarian carcinoma OVCAR-8, and Burkitt's lymphoma CA46 cell lines. RAF1 depletion by siRNAs attenuated LSA-induced PCC in HCT-116 and OVCAR-8 cells. Furthermore, mouse embryonic fibroblasts (MEF) with homozygous deletion in Raf1, but not deletion in the related kinase Braf, were resistant to LSA-induced PCC. Complementation of Raf1-/- MEFs with wild-type human RAF1, but not with kinase-dead RAF1 mutant, restored LSA-induced PCC. Finally, the Raf inhibitor sorafenib, but not the MEK inhibitor AZD6244, effectively suppressed LSA-induced PCC. Our findings implicate a previously unknown, MAPK-independent role of RAF1 in chromatin condensation and potent activation of this pathway by LSA.
authors.affiliation: Developmental Therapeutics Branch and Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute (CCR-NCI), NIH, Bethesda, MD 20892-9760, USA. rozenn.josse@gmail.com.
authors.forename: Rozenn
authors.initials: R
authors.lastname: Jossé
authors.affiliation: Developmental Therapeutics Branch and Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute (CCR-NCI), NIH, Bethesda, MD 20892-9760, USA. yongwei.zhang@georgetown.edu.
authors.forename: Yong-Wei
authors.initials: YW
authors.lastname: Zhang
authors.affiliation: Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute (CCR-NCI), NIH, Bethesda, MD 20892-9760, USA. valentin_giroux@hotmail.com.
authors.forename: Valentin
authors.initials: V
authors.lastname: Giroux
authors.affiliation: Departments of Chemistry and Medicinal Chemistry, Purdue University; West Lafayette, IN 47907, USA. akghosh@purdue.edu.
authors.forename: Arun K
authors.initials: AK
authors.lastname: Ghosh
authors.affiliation: Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute (CCR-NCI), NIH, Bethesda, MD 20892-9760, USA. ji.luo@nih.gov.
authors.forename: Ji
authors.initials: J
authors.lastname: Luo
authors.affiliation: Developmental Therapeutics Branch and Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute (CCR-NCI), NIH, Bethesda, MD 20892-9760, USA. pommier@nih.gov.
authors.forename: Yves
authors.initials: Y
authors.lastname: Pommier
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000302:isolation & purification / Q000494:pharmacology*; D001562:Benzimidazoles / Q000494:pharmacology; D045744:Cell Line, Tumor; D002843:Chromatin / Q000187:drug effects / Q000378:metabolism; D002875:Chromosomes / Q000187:drug effects / Q000378:metabolism; D004622:Embryo, Mammalian; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D018942:Macrolides / Q000302:isolation & purification / Q000494:pharmacology*; D051379:Mice; D009536:Niacinamide / Q000031:analogs & derivatives / Q000494:pharmacology; D010671:Phenylurea Compounds / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011161:Porifera / Q000737:chemistry*; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D000077157:Sorafenib
chemical_list: C517975:AZD 6244; D000970:Antineoplastic Agents; D001562:Benzimidazoles; D002843:Chromatin; D018942:Macrolides; D010671:Phenylurea Compounds; C450700:lasonolide A; D009536:Niacinamide; D000077157:Sorafenib; D019908:Proto-Oncogene Proteins c-raf
keywords: c-raf; chromosome condensation; lasonolide A; myosin phosphatase; shRNA