Host cell factors controlling vimentin organization in the Xenopus oocyte.

Abstract

Abstract. To study vimentin filament organization in vivo we injected Xenopus oocytes, which have no significant vimentin system of their own, with in vitro-synthesized RNAs encoding Xenopus vimentins.Exogenous vimentins were localized primarily to the cytoplasmic surface of the nucleus and to the subplasma membrane "cortex." In the cortex of the animal hemisphere, wild-type vimentin forms punctate structures and short filaments. In contrast, long anastomosing vimentin filaments are formed in the vegetal hemisphere cortex. This asymmetry in the organization of exogenous vimentin is similar to that of the endogenous keratin system (Klymkowsky, M. W., L. A. vimentin filaments remain intact and vimentin is hyperphosphorylated. To examine the role of MPF kinase in the M-phase reorganization of vimentin we deleted the conserved proline of vimentin's single MPF-kinase site; this mutation had no apparent effect on the prophase or M-phase behavior of vimentin. In contrast, deletion of amino acids 19-68 or 18-61 of the NH2-terminal "head" domain produced proteins that formed extended filaments in the animal hemisphere of the prophase oocyte. We suggest that the animal hemisphere cortex of the prophase oocyte contains a factor that actively suppresses the formation of extended vimentin filaments through a direct interaction with vimentin's head domain. During maturation this "suppressor of extended filaments" appears to be inactivated, leading to the formation of an extended vimentin filament system.

Nuclear pore disassembly from endoplasmic reticulum membranes promotes Ca2+signalling competency

“…These data imply that MPF activation parallels the timeframe of AL disappearance, and that the kinase is ‘globally’ active in cells where AL remodelling is occurring, but do not demonstrate that MPF is active in the local vicinity of AL prior to NPC dissociation, as expected if this kinase was a key trigger for NPC dissociation in vivo . Therefore, to monitor endogenous MPF activity, we expressed a GFP‐tagged vimentin, a substrate for MPF that targets the oocyte cortex (Dent et al 1992). Vimentin is phosphorylated by MPF at a site in the NH 2 ‐terminus (Ser‐55), the phosphorylation status of which can be resolved using phospho‐specific antibodies.…”

Confocal microscopy and 3‐D reconstruction of the cytoskeleton of Xenopus oocytes

“…Although the presence of vimentin in Xenopus oocytes remains controversial (Dent et al, 1992;Franz et al, 1983;Herrmann et al, 1989), the presence of a complex network of KFs is indisputable. Conventional microscopy of whole-mounted or sectioned stage VI Xenopus oocytes stained with antibodies to keratins revealed a complex, anastomosing network of KFs in the vegetal cortex, radial networks of cytoplasmic KFs, and a network of KFs surrounding the GV (Franz et al, 1983;Godsave et al, 1984b;Klymkowsky et al, 1987;Ryabova et al, 1993;Torpey et al, 1992b).…”

The Sox axis, Nodal signaling, and germ layer specification

“…laevis . Our interests (Klymkowsky et al, 1987, 1991; Dent et al, 1992; Bachant, 1993; Bachant and Klymkowsky, 1996) intersected with his (Kloc et al, 2005), with respect to the asymmetric organization and dynamics of the oocyte's keratin‐type intermediate filament network. It was therefore a pleasant surprise to find that our work on Soxs has brought us back to maternal asymmetries, and we were beginning a project with Larry and his long‐time collaborator Malgorzata Kloc on the localization of Sox7 mRNA during oogenesis.…”