title: Dephosphorylation of MAP2D enhances its binding to vimentin in preovulatory ovarian granulosa cells.
abstract: Preovulatory granulosa cells express the low-molecular-mass MAP2D variant of microtubule-associated protein 2 (MAP2). Activation of the luteinizing hormone choriogonadotropin receptor by human choriogonadotropin (hCG) promotes dephosphorylation of MAP2D on Thr256 and Thr259. We sought to evaluate the association of MAP2D with the cytoskeleton, and the effect of hCG on this association. MAP2D partially colocalized, as assessed by confocal immunofluorescence microscopy, with the vimentin intermediate filament and microtubule cytoskeletons in naive cells. In vitro binding studies showed that MAP2D bound directly to vimentin and β-tubulin. Phosphorylation of recombinant MAP2D on Thr256 and Thr259, which mimics the phosphorylation status of MAP2D in naive cells, reduces binding of MAP2D to vimentin and tubulin by two- and three-fold, respectively. PKA-dependent phosphorylation of vimentin (Ser32 and Ser38) promoted binding of vimentin to MAP2D and increased contraction of granulosa cells with reorganization of vimentin filaments and MAP2D from the periphery into a thickened layer surrounding the nucleus and into prominent cellular extensions. Chemical disruption of vimentin filament organization increased progesterone production. Taken together, these results suggest that hCG-stimulated dephosphorylation of MAP2D at Thr256 and Thr259, phosphorylation of vimentin at Ser38 and Ser72, and the resulting enhanced binding of MAP2D to vimentin might contribute to the progesterone synthetic response required for ovulation.
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA.
authors.forename: Maxfield P
authors.identifier: 0000-0002-6343-0150
authors.initials: MP
authors.lastname: Flynn
authors.affiliation: Department of Medicine, Oregon Health and Sciences University and VA Portland Health Care System, Portland, OR 97239, USA.
authors.forename: Sarah E
authors.identifier: 0000-0002-1866-9074
authors.initials: SE
authors.lastname: Fiedler
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA School of Molecular Biosciences, Washington State University, Pullman, WA 99164, USA.
authors.forename: Amelia B
authors.identifier: 0000-0002-4413-4217
authors.initials: AB
authors.lastname: Karlsson
authors.affiliation: Department of Medicine, Oregon Health and Sciences University and VA Portland Health Care System, Portland, OR 97239, USA.
authors.forename: Daniel W
authors.identifier: 0000-0003-1954-2478
authors.initials: DW
authors.lastname: Carr
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA.
authors.forename: Evelyn T
authors.identifier: 0000-0001-9072-1095
authors.initials: ET
authors.lastname: Maizels
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA School of Molecular Biosciences, Washington State University, Pullman, WA 99164, USA mehd@wsu.edu.
authors.forename: Mary
authors.identifier: 0000-0003-4640-1814
authors.initials: M
authors.lastname: Hunzicker-Dunn
mesh_terms: D000818:Animals; D006063:Chorionic Gonadotropin / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D005260:Female; D005455:Fluorescent Antibody Technique; D000071679:Glycogen Synthase Kinase 3 beta / Q000378:metabolism; D006107:Granulosa Cells / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D007382:Intermediate Filaments / Q000187:drug effects / Q000378:metabolism; D018613:Microscopy, Confocal; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism; D008875:Middle Aged; D050505:Mutant Proteins / Q000378:metabolism; D010060:Ovulation / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D010769:Phosphothreonine / Q000378:metabolism; D011374:Progesterone / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000378:metabolism; D012995:Solubility; D014746:Vimentin / Q000378:metabolism*
chemical_list: D006063:Chorionic Gonadotropin; C489787:MAP2 protein, rat; D008869:Microtubule-Associated Proteins; D050505:Mutant Proteins; D011994:Recombinant Proteins; D014746:Vimentin; D010769:Phosphothreonine; D011374:Progesterone; D000071679:Glycogen Synthase Kinase 3 beta; D017868:Cyclic AMP-Dependent Protein Kinases
keywords: Choriogonadotropin receptor; Granulosa cell; Luteinizing hormone receptor; MAP2D; PKA; Vimentin