title: Readthrough acetylcholinesterase (AChE-R) and regulated necrosis: pharmacological targets for the regulation of ovarian functions?
abstract: Proliferation, differentiation and death of ovarian cells ensure orderly functioning of the female gonad during the reproductive phase, which ultimately ends with menopause in women. These processes are regulated by several mechanisms, including local signaling via neurotransmitters. Previous studies showed that ovarian non-neuronal endocrine cells produce acetylcholine (ACh), which likely acts as a trophic factor within the ovarian follicle and the corpus luteum via muscarinic ACh receptors. How its actions are restricted was unknown. We identified enzymatically active acetylcholinesterase (AChE) in human ovarian follicular fluid as a product of human granulosa cells. AChE breaks down ACh and thereby attenuates its trophic functions. Blockage of AChE by huperzine A increased the trophic actions as seen in granulosa cells studies. Among ovarian AChE variants, the readthrough isoform AChE-R was identified, which has further, non-enzymatic roles. AChE-R was found in follicular fluid, granulosa and theca cells, as well as luteal cells, implying that such functions occur in vivo. A synthetic AChE-R peptide (ARP) was used to explore such actions and induced in primary, cultured human granulosa cells a caspase-independent form of cell death with a distinct balloon-like morphology and the release of lactate dehydrogenase. The RIPK1 inhibitor necrostatin-1 and the MLKL-blocker necrosulfonamide significantly reduced this form of cell death. Thus a novel non-enzymatic function of AChE-R is to stimulate RIPK1/MLKL-dependent regulated necrosis (necroptosis). The latter complements a cholinergic system in the ovary, which determines life and death of ovarian cells. Necroptosis likely occurs in the primate ovary, as granulosa and luteal cells were immunopositive for phospho-MLKL, and hence necroptosis may contribute to follicular atresia and luteolysis. The results suggest that interference with the enzymatic activities of AChE and/or interference with necroptosis may be novel approaches to influence ovarian functions.
authors.affiliation: Anatomy III - Cell Biology, Ludwig-Maximilian-University (LMU), Munich, Germany.
authors.forename: J
authors.initials: J
authors.lastname: Blohberger
authors.affiliation: Neurobiology, Department Biology II, Ludwig-Maximilian-University (LMU), Munich, Germany.
authors.forename: L
authors.initials: L
authors.lastname: Kunz
authors.affiliation: Anatomy III - Cell Biology, Ludwig-Maximilian-University (LMU), Munich, Germany.
authors.forename: D
authors.identifier: 0000000155412799
authors.initials: D
authors.lastname: Einwang
authors.affiliation: ART, Bogenhausen, Munich, Germany.
authors.forename: U
authors.initials: U
authors.lastname: Berg
authors.affiliation: ART, Bogenhausen, Munich, Germany.
authors.forename: D
authors.initials: D
authors.lastname: Berg
authors.affiliation: Oregon National Primate Research Center/OHSU, Beaverton, OR, USA.
authors.forename: S R
authors.initials: SR
authors.lastname: Ojeda
authors.affiliation: Oregon National Primate Research Center/OHSU, Beaverton, OR, USA.
authors.forename: G A
authors.initials: GA
authors.lastname: Dissen
authors.affiliation: Laboratory for Functional Genome Analysis LAFUGA, Gene Center, Ludwig-Maximilian-University (LMU), Munich, Germany.
authors.forename: T
authors.initials: T
authors.lastname: Fröhlich
authors.affiliation: Laboratory for Functional Genome Analysis LAFUGA, Gene Center, Ludwig-Maximilian-University (LMU), Munich, Germany.
authors.forename: G J
authors.initials: GJ
authors.lastname: Arnold
authors.affiliation: Department of Biological Chemistry and the Edmond and Lily Safra Center of Brain Science, The Hebrew University of Jerusalem, The Edmond J. Safra Campus, Givat Ram, Jerusalem, Israel.
authors.forename: H
authors.initials: H
authors.lastname: Soreq
authors.affiliation: Laboratorio de Neurobioquimica, Facultad de Ciencias Quimicas y Farmaceuticas, Universidad de Chile, Santiago, Chile.
authors.forename: H
authors.initials: H
authors.lastname: Lara
authors.affiliation: Anatomy III - Cell Biology, Ludwig-Maximilian-University (LMU), Munich, Germany.
authors.forename: A
authors.initials: A
authors.lastname: Mayerhofer
mesh_terms: D000109:Acetylcholine / Q000378:metabolism; D000110:Acetylcholinesterase / Q000096:biosynthesis* / Q000378:metabolism; D000178:Acrylamides / Q000008:administration & dosage; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics; D005260:Female; D006107:Granulosa Cells / Q000187:drug effects / Q000201:enzymology*; D006801:Humans; D007093:Imidazoles / Q000008:administration & dosage; D007211:Indoles / Q000008:administration & dosage; D006080:Ovarian Follicle / Q000254:growth & development / Q000378:metabolism*; D061251:Primary Cell Culture; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D013449:Sulfonamides / Q000008:administration & dosage
chemical_list: D000178:Acrylamides; D007093:Imidazoles; D007211:Indoles; C570695:N-(4-(N-(3-methoxypyrazin-2-yl)sulfamoyl)phenyl)-3-(5-nitrothiophene-2-yl)acrylamide; D013449:Sulfonamides; C507699:necrostatin-1; C570693:MLKL protein, human; D011494:Protein Kinases; C506249:RIPK1 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; D000110:Acetylcholinesterase; D000109:Acetylcholine