title: cFLIP regulates death receptor-mediated apoptosis in an ovarian granulosa cell line by inhibiting procaspase-8 cleavage.
abstract: More than 99% of follicles in mammalian ovaries undergo atresia, but the mechanisms regulating the strict selection process are still unclear. Granulosa cell apoptosis is considered the trigger of follicular atresia, which occurs in advance of the death of an oocyte. Cellular FLICE-like inhibitory protein (cFLIP), a homologue of procaspase-8 (also called FLICE), is an intracellular anti-apoptotic protein. It is expressed in granulosa cells of porcine ovaries, where its levels decreases during follicular atresia. We hypothesized that cFLIP regulates granulosa cell apoptosis by acting as a pro-survival factor. In the present study, to further reveal the function of cFLIP in granulosa cells, we examined the anti-apoptotic mechanism of cFLIP using KGN, a human granulosa tumor cell line. Fas-mediated apoptosis was induced by co-treatment with anti-Fas antibody (CH-11), which acts as an agonist of Fas-ligand, and cycloheximide (CHX). When cFLIP was stably expressed in KGN cells following transfection of an expression vector, the Fas-mediated apoptosis was inhibited. Suppression of cFLIP by small interfering RNA (siRNA) spontaneously induced cell death. Silencing of cFLIP promoted cleavage of procaspase-8, and the cell death caused by cFLIP siRNA was completely blocked by a caspase-8 inhibitor (Z-IETD-FMK), indicating that cFLIP regulates apoptosis in KGN cells by inhibiting cleavage of procaspase-8. In conclusion, cFLIP is an essential pro-survival factor for granulosa cells, and it prevents granulosa cell apoptosis by inhibiting procaspase-8 activation.
authors.affiliation: Research Unit for Animal Life Sciences, Animal Resource Science Center, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Ibaraki-Kasama, Japan. Fuko_Minehata@nts.toray.co.jp
authors.forename: Fuko
authors.initials: F
authors.lastname: Matsuda
authors.forename: Naoko
authors.initials: N
authors.lastname: Inoue
authors.forename: Yasufumi
authors.initials: Y
authors.lastname: Goto
authors.forename: Akihisa
authors.initials: A
authors.lastname: Maeda
authors.forename: Yuan
authors.initials: Y
authors.lastname: Cheng
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Sakamaki
authors.forename: Noboru
authors.initials: N
authors.lastname: Manabe
mesh_terms: D017209:Apoptosis / Q000187:drug effects / Q000235:genetics*; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D053181:Caspase 8 / Q000378:metabolism; D061945:Caspase Inhibitors; D002460:Cell Line; D004789:Enzyme Activation / Q000187:drug effects / Q000235:genetics; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D006107:Granulosa Cells / Q000378:metabolism / Q000502:physiology; D006801:Humans; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D034741:RNA, Small Interfering / Q000494:pharmacology; D053218:Receptors, Death Domain / Q000235:genetics / Q000502:physiology; D019014:fas Receptor / Q000502:physiology
chemical_list: D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; C506386:CFLAR protein, human; D061945:Caspase Inhibitors; D034741:RNA, Small Interfering; D053218:Receptors, Death Domain; D019014:fas Receptor; D053181:Caspase 8