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Intranuclear membranes induced by lipidated proteins are derived from the nuclear envelope.

ABSTRACT: Association of nuclear lamins with the inner nuclear membrane (INM) is mediated by lipid modifications: either by C-terminal isoprenylation or N-terminal myristoylation. Overexpression of lamins or other lipidated nuclear proteins induces the formation of intranuclear membrane-like arrays. Lamin-induced intranuclear array formation has been observed in Xenopus oocytes as well as in mammalian tissue culture cells. With the use of a membrane-specific fluorescence dye we show here that these arrays are made up of typical lipid membranes. While continuity between these intranuclear membranes and the INM has not been observed so far the presence of integral as well as luminal marker proteins of the endoplasmic reticulum (ER) indicates that these membranes are derived from the nuclear membrane/ER compartment. Earlier studies demonstrated that overexpression of integral membrane proteins of the INM can induce formation of intranuclear membranes, which bud from the INM. Integral membrane proteins reach the INM via the pore membranes while lipidated proteins are imported into the nucleoplasm via the classical NLS pathway where they interact with the INM via their lipid moieties. Together with the previously published data our results show that the formation of intranuclear membranes follows similar routes irrespective of whether the proteins triggering membrane formation are integral membrane or lipidated proteins.

SUBMITTER: Linde N

PROVIDER: S-EPMC3027043 | biostudies-literature | 2010 Jul-Aug

REPOSITORIES: biostudies-literature

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Intranuclear membranes induced by lipidated proteins are derived from the nuclear envelope.

Linde Nina N Stick Reimer R

Nucleus (Austin, Tex.) 20100428 4

Association of nuclear lamins with the inner nuclear membrane (INM) is mediated by lipid modifications: either by C-terminal isoprenylation or N-terminal myristoylation. Overexpression of lamins or other lipidated nuclear proteins induces the formation of intranuclear membrane-like arrays. Lamin-induced intranuclear array formation has been observed in Xenopus oocytes as well as in mammalian tissue culture cells. With the use of a membrane-specific fluorescence dye we show here that these arrays ...[more]

PMID: 21327083

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Project description:The nuclear envelope (NE) consists of two concentric nuclear membranes separated by the lumen, an ∼40-nm-wide fluid layer. NE proteins are implicated in important cellular processes ranging from gene expression to nuclear positioning. Although recent progress has been achieved in quantifying the assembly states of NE proteins in their native environment with fluorescence fluctuation spectroscopy, these studies raised questions regarding the association of NE proteins with nuclear membranes during the assembly process. Monitoring the interaction of proteins with membranes is important because the binding event is often associated with conformational changes that are critical to cellular signaling pathways. Unfortunately, the close physical proximity of both membranes poses a severe experimental challenge in distinguishing luminal and membrane-associated NE proteins. This study seeks to address this problem by introducing new, to our knowledge, fluorescence-based assays that overcome the restrictions imposed by the NE environment. We found that luminal proteins violate the Stokes-Einstein relation, which eliminates a straightforward use of protein mobility as a marker of membrane association within the NE. However, a surprising anomaly in the temperature-dependent mobility of luminal proteins was observed, which was developed into an assay for distinguishing between soluble and membrane-bound NE proteins. We further introduced a second independent tool for distinguishing both protein populations by harnessing the previously reported undulations of the nuclear membranes. These membrane undulations introduce local volume changes that produce an additional fluorescence fluctuation signal for luminal, but not for membrane-bound, proteins. After testing both methods using simple model systems, we apply the two assays to investigate a previously proposed model of membrane association for the luminal domain of SUN2, a constituent protein of the linker of nucleoskeleton and cytoskeleton complex. Finally, we investigate the effect of C- and N-terminal tagging of the luminal ATPase torsinA on its ability to associate with nuclear membranes.

Dataset Information

Intranuclear membranes induced by lipidated proteins are derived from the nuclear envelope.

Publications

Intranuclear membranes induced by lipidated proteins are derived from the nuclear envelope.

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