scholarly journals Requirement for Lamin B Receptor and Its Regulation by Importin β and Phosphorylation in Nuclear Envelope Assembly during Mitotic Exit

2010 ◽  
Vol 285 (43) ◽  
pp. 33281-33293 ◽  
Author(s):  
Xuelong Lu ◽  
Yang Shi ◽  
Quanlong Lu ◽  
Yan Ma ◽  
Jia Luo ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Mitotic Exit ◽  
Lamin B ◽  
Lamin B Receptor ◽  
Nuclear Envelope Assembly

scholarly journals Temporal control of nuclear envelope assembly by phosphorylation of lamin B receptor

2011 ◽  
Vol 22 (18) ◽  
pp. 3306-3317 ◽  
Author(s):  
Li-Chuan Tseng ◽  
Rey-Huei Chen
Keyword(s):  
Membrane Protein ◽  
Nuclear Envelope ◽  
Nuclear Membrane ◽  
Chromatin Binding ◽  
Temporal Control ◽  
Lamin B ◽  
Lamin B Receptor ◽  
Nuclear Envelope Assembly ◽  
Nuclear Membrane Protein

The nuclear envelope of metazoans disassembles during mitosis and reforms in late anaphase after sister chromatids have well separated. The coordination of these mitotic events is important for genome stability, yet the temporal control of nuclear envelope reassembly is unknown. Although the steps of nuclear formation have been extensively studied in vitro using the reconstitution system from egg extracts, the temporal control can only be studied in vivo. Here, we use time-lapse microscopy to investigate this process in living HeLa cells. We demonstrate that Cdk1 activity prevents premature nuclear envelope assembly and that phosphorylation of the inner nuclear membrane protein lamin B receptor (LBR) by Cdk1 contributes to the temporal control. We further identify a region in the nucleoplasmic domain of LBR that inhibits premature chromatin binding of the protein. We propose that this inhibitory effect is partly mediated by Cdk1 phosphorylation. Furthermore, we show that the reduced chromatin-binding ability of LBR together with Aurora B activity contributes to nuclear envelope breakdown. Our studies reveal for the first time a mechanism that controls the timing of nuclear envelope reassembly through modification of an integral nuclear membrane protein.

Alterations of nuclear envelope and chromatin organization in mandibuloacral dysplasia, a rare form of laminopathy

2005 ◽  
Vol 23 (2) ◽  
pp. 150-158 ◽  
Author(s):  
Ilaria Filesi ◽  
Francesca Gullotta ◽  
Giovanna Lattanzi ◽  
Maria Rosaria D'Apice ◽  
Cristina Capanni ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Protein A ◽  
Nuclear Architecture ◽  
Mendelian Inheritance ◽  
Premature Aging ◽  
Lamin A ◽  
Lamin B ◽  
Lamin B Receptor ◽  
Epigenetic Events ◽  
Mandibuloacral Dysplasia

Autosomal recessive mandibuloacral dysplasia [mandibuloacral dysplasia type A (MADA); Online Mendelian Inheritance in Man (OMIM) no. 248370 ] is caused by a mutation in LMNA encoding lamin A/C. Here we show that this mutation causes accumulation of the lamin A precursor protein, a marked alteration of the nuclear architecture and, hence, chromatin disorganization. Heterochromatin domains are altered or completely lost in MADA nuclei, consistent with the finding that heterochromatin-associated protein HP1β and histone H3 methylated at lysine 9 and their nuclear envelope partner protein lamin B receptor (LBR) are delocalized and solubilized. Both accumulation of lamin A precursor and chromatin defects become more severe in older patients. These results strongly suggest that altered chromatin remodeling is a key event in the cascade of epigenetic events causing MADA and could be related to the premature-aging phenotype.

BAF is required for emerin assembly into the reforming nuclear envelope

2001 ◽  
Vol 114 (24) ◽  
pp. 4575-4585 ◽  
Author(s):  
Tokuko Haraguchi ◽  
Takako Koujin ◽  
Miriam Segura-Totten ◽  
Kenneth K. Lee ◽  
Yosuke Matsuoka ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Hela Cells ◽  
Core Region ◽  
Lamin B ◽  
The Core ◽  
Double Stranded Dna ◽  
Recessive Form ◽  
Nuclear Envelope Assembly

Mutations in emerin cause the X-linked recessive form of Emery-Dreifuss muscular dystrophy (EDMD). Emerin localizes at the inner membrane of the nuclear envelope (NE) during interphase, and diffuses into the ER when the NE disassembles during mitosis. We analyzed the recruitment of wildtype and mutant GFP-tagged emerin proteins during nuclear envelope assembly in living HeLa cells. During telophase, emerin accumulates briefly at the ‘core’ region of telophase chromosomes, and later distributes over the entire nuclear rim. Barrier-to-autointegration factor (BAF), a protein that binds nonspecifically to double-stranded DNA in vitro, co-localized with emerin at the ‘core’ region of chromosomes during telophase. An emerin mutant defective for binding to BAF in vitro failed to localize at the ‘core’ in vivo, and subsequently failed to localize at the reformed NE. In HeLa cells that expressed BAF mutant G25E, which did not show ‘core’ localization, the endogenous emerin proteins failed to localize at the ‘core’ region during telophase, and did not assemble into the NE during the subsequent interphase. BAF mutant G25E also dominantly dislocalized LAP2β and lamin A from the NE, but had no effect on the localization of lamin B. We conclude that BAF is required for the assembly of emerin and A-type lamins at the reforming NE during telophase, and may mediate their stability in the subsequent interphase.

scholarly journals Induction of a Massive Endoplasmic Reticulum and Perinuclear Space Expansion by Expression of Lamin B Receptor Mutants and the Related Sterol Reductases TM7SF2 and DHCR7

2010 ◽  
Vol 21 (2) ◽  
pp. 354-368 ◽  
Author(s):  
Monika Zwerger ◽  
Thorsten Kolb ◽  
Karsten Richter ◽  
Iakowos Karakesisoglou ◽  
Harald Herrmann
Keyword(s):  
Endoplasmic Reticulum ◽  
Nuclear Envelope ◽  
Cell Lines ◽  
Nuclear Membrane ◽  
Cultured Cells ◽  
Membrane Separation ◽  
Reductase Activity ◽  
Nuclear Pore ◽  
Lamin B ◽  
Lamin B Receptor

Lamin B receptor (LBR) is an inner nuclear membrane protein involved in tethering the nuclear lamina and the underlying chromatin to the nuclear envelope. In addition, LBR exhibits sterol reductase activity. Mutations in the LBR gene cause two different human diseases: Pelger-Huët anomaly and Greenberg skeletal dysplasia, a severe chrondrodystrophy causing embryonic death. Our study aimed at investigating the effect of five LBR disease mutants on human cultured cells. Three of the tested LBR mutants caused a massive compaction of chromatin coincidental with the formation of a large nucleus-associated vacuole (NAV) in several human cultured cell lines. Live cell imaging and electron microscopy revealed that this structure was generated by the separation of the inner and outer nuclear membrane. During NAV formation, nuclear pore complexes and components of the linker of nucleoskeleton and cytoskeleton complex were lost in areas of membrane separation. Concomitantly, a large number of smaller vacuoles formed throughout the cytoplasm. Notably, forced expression of the two structurally related sterol reductases transmembrane 7 superfamily member 2 and 7-dehydrocholesterol reductase caused, even in their wild-type form, a comparable phenotype in susceptible cell lines. Hence, LBR mutant variants and sterol reductases can severely interfere with the regular organization of the nuclear envelope and the endoplasmic reticulum.

scholarly journals Cytomegaloviral proteins that associate with the nuclear lamina: components of a postulated nuclear egress complex

2009 ◽  
Vol 90 (3) ◽  
pp. 579-590 ◽  
Author(s):  
Jens Milbradt ◽  
Sabrina Auerochs ◽  
Heinrich Sticht ◽  
Manfred Marschall
Keyword(s):  
Nuclear Envelope ◽  
Transmembrane Protein ◽  
Nuclear Lamina ◽  
Cellular Proteins ◽  
Lamin B ◽  
Nuclear Egress ◽  
Lamin B Receptor ◽  
Individual Interaction ◽  
Sequence Elements ◽  
Protein Recruitment

The nuclear egress of cytomegaloviral capsids traversing the nuclear envelope is dependent on a locally restricted destabilization of the rigid nuclear lamina. It has been suggested that the multi-component nuclear egress complex (NEC) that is formed is comprised of both viral and cellular proteins which act to recruit lamin-phosphorylating protein kinases. Recently, we reported that the lamina-associated human cytomegalovirus-encoded proteins pUL50 and pUL53, conserved among herpesviruses, interact with each other and recruit protein kinase C (PKC) to the nuclear envelope in transfected cells. The multiple interactions of the transmembrane protein pUL50 with pUL53, PKC and cellular PKC-binding protein p32, appear crucial to the formation of the NEC. In this study, we mapped individual interaction sequence elements of pUL50 by coimmunoprecipitation analysis of deletion mutants and yeast two-hybrid studies. Amino acids 1–250 were shown to be responsible for interaction with pUL53, 100–280 for PKC and 100–358 for p32. Interestingly, p32 specifically interacted with multiple NEC components, including the kinases PKC and pUL97, thus possibly acting as an adaptor for protein recruitment to the lamin B receptor. Notably, p32 was the only protein that interacted with the lamin B receptor. Immunofluorescence studies visualized the colocalization of NEC components at the nuclear rim in coexpression studies. The data imply that a tight interaction between at least six viral and cellular proteins leads to the formation of a postulated multi-protein complex required for nuclear egress.

scholarly journals Lamin A, lamin B, and lamin B receptor analogues in yeast.

1989 ◽  
Vol 108 (6) ◽  
pp. 2069-2082 ◽  
Author(s):  
S D Georgatos ◽  
I Maroulakou ◽  
G Blobel
Keyword(s):  
Nuclear Envelope ◽  
Peptide Mapping ◽  
Polyclonal Antibodies ◽  
Structural Similarity ◽  
Integral Membrane Protein ◽  
Lamin A ◽  
Nuclear Fraction ◽  
Yeast Saccharomyces Cerevisiae ◽  
Lamin B ◽  
Lamin B Receptor

Previous studies have shown that turkey erythrocyte lamin B is anchored to the nuclear envelope via a 58-kD integral membrane protein termed p58 or lamin B receptor (Worman H. J., J. Yuan, G. Blobel, and S. D. Georgatos. 1988. Proc. Natl. Acad. Sci. USA. 85:8531-8534). We now identify a p58 analogue in the yeast Saccharomyces cerevisiae. Turkey erythrocyte lamin B binds to yeast urea-extracted nuclear envelopes with high affinity, associating predominantly with a 58-kD polypeptide. This yeast polypeptide is recognized by polyclonal antibodies against turkey p58, partitions entirely with the nuclear fraction, remains membrane bound after urea extraction of the nuclear envelopes, and is structurally similar to turkey p58 by peptide mapping criteria. Using polyclonal antibodies against turkey erythrocyte lamins A and B, we also identify two yeast lamin forms. The yeast lamin B analogue has a molecular mass of 66 kD and is structurally related to erythrocyte lamin B. Moreover, the yeast lamin B analogue partitions exclusively with the nuclear envelope fraction, is quantitatively removed from the envelopes by urea extraction, and binds to turkey lamin A and vimentin. As many higher eukaryotic lamin B forms, the yeast analogue is chemically heterogeneous comprising two serologically related species with different charge characteristics. Antibodies against turkey lamin A detect a 74-kD yeast protein, slightly larger than the turkey lamin A. It is more abundant than the yeast lamin B analogue and partitions between a soluble cytoplasmic fraction and a nuclear envelope fraction. The yeast lamin A analogue can be extracted from the nuclear envelope by urea, shows structural similarity to turkey and rat lamin A, and binds to isolated turkey lamin B. These data indicate that analogues of typical nuclear lamina components (lamins A and B, as well as lamin B receptor) are present in yeast and behave as their vertebrate counterparts.

Lamin B receptor-mediated chromatin tethering to the nuclear envelope is detrimental to the xenopus blastula

2020 ◽  
Author(s):  
Haruka Oda ◽  
Satsuki Kato ◽  
Keita Ohsumi ◽  
Mari Iwabuchi
Keyword(s):  
Nuclear Envelope ◽  
Mitotic Spindles ◽  
Filamentous Actin ◽  
Blastula Stage ◽  
Lamin B ◽  
Lamin B Receptor ◽  
Specific Manner ◽  
Egg Extracts ◽  
Xenopus Egg ◽  
Xenopus Egg Extracts

Abstract In the nucleus of eukaryotic cells, chromatin is tethered to the nuclear envelope (NE), wherein inner nuclear membrane proteins (INMPs) play major roles. However, in Xenopus blastula, chromatin tethering to the NE depends on nuclear filamentous actin that develops in a blastula-specific manner. To investigate whether chromatin tethering operates in the blastula through INMPs, we experimentally introduced INMPs into Xenopus egg extracts that recapitulate nuclear formation in fertilized eggs. When expressed in extracts in which polymerization of actin is inhibited, only lamin B receptor (LBR), among the five INMPs tested, tethered chromatin to the NE, depending on its N2 and N3 domains responsible for chromatin-protein binding. N2-3-deleted LBR did not tether chromatin, although it was localized in the nuclei. We subsequently found that the LBR level was very low in the Xenopus blastula but was elevated after the blastula stage. When the LBR level was precociously elevated in the blastula by injecting LBR mRNA, it induced alterations in nuclear laminar architecture and nuclear morphology, and caused DNA damage and abnormal mitotic spindles, depending on the N2-3 domains. These results suggest that LBR-mediated chromatin tethering is circumvented in the Xenopus blastula, as it is detrimental to embryonic development.

Transcriptional Regulation of the Nuclear Envelope Lamin B Receptor: New Roles for GA Binding Protein (GABP) in Neutrophil Development.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1464-1464
Author(s):  
Rahul Garhwal ◽  
Alan G. Rosmarin ◽  
Stephanie Halene ◽  
Peter Gaines
Keyword(s):  
Nuclear Envelope ◽  
Binding Sites ◽  
Envelope Protein ◽  
Gene Activation ◽  
Gene Promoter ◽  
Functional Genes ◽  
Lamin B ◽  
Lamin B Receptor ◽  
Ets Family ◽  
Ga Binding Protein

Abstract Abstract 1464 Poster Board I-487 The essential roles that neutrophils play in innate immune responses require that these phagocytes rapidly migrate to sites of infection, adhere to and escape capillary epithelium, and then engulf and destroy invading pathogens. The capacity of neutrophils to perform these complicated functions is largely dependent on the actions of multiple hematopoietic transcription factors that coordinate the expression of critical functional genes during neutrophil development. These transcription factors include CCAAT/enhancer binding proteins C/EBPα and C/EBPε, the ETS family protein PU.1, and the GA binding protein, GABP, which is also an ETS transcription factor that acts as an obligate heterotetramer comprised of GABPα and GABPβ. In addition to the activation of functional genes, neutrophil progenitors in the bone marrow undergo profound morphologic changes that include the formation of lobulated nuclei. Although it is still unclear as to the precise purpose of nuclear lobulation, we and others have demonstrated that the nuclear envelope protein called the lamin B receptor (LBR) is essential to neutrophil nuclear lobulation and that loss of Lbr expression in mouse neutrophils leads to decreased functional responses, including a reduced respiratory burst and abnormal chemotaxis. We also have shown that Lbr gene expression is upregulated during neutrophil development, indicating that the transcriptional control of Lbr expression may play a critical role in both neutrophil morphologic maturation and function. To identify the transcriptional regulators that control Lbr activation during neutrophil development, we have isolated the promoter of the mouse Lbr gene and have assessed the roles that different regulators of neutrophil development play on Lbr activation. Previous studies demonstrated that C/EBPε directly activates the Lbr promoter, but we found that myeloid C/EBPε-/- cells exhibit normal Lbr expression. We therefore focused on identifying alternative regulators that may control Lbr gene activation. We first identified two putative ets binding sites in the Lbr gene promoter that are located near two previously identified C/EBP binding sites. We then focused on analyzing transcriptional activities of two ETS family members known to play roles in myeloid gene activation, PU.1 and GABP, on Lbr promoter sequences that contain these ets binding sites. Using promoter expression constructs that contain different sized regions of the Lbr gene promoter and Cos cell transfections, we confirm that C/EBPε alone does indeed drive Lbr expression, but that either C/EBPα or PU.1 alone fails to activate the Lbr promoter. In contrast, expression of GABP, via co-expression of GABPα and GABPβ, transcriptionally activates the Lbr promoter to levels observed with C/EBPε alone. Interestingly, GABP activates a short Lbr promoter that contains only one ets binding site but that lacks C/EBPε binding sites, indicating that a single ets site is sufficient for GABP to drive Lbr expression in the absence of C/EBPε. Furthermore, co-expression of GABP plus PU.1 significantly increased activation of the Lbr promoter to levels above that observed for either GABP alone or C/EBPε alone. This result indicates that GABP and PU.1 synergistically activate the Lbr promoter. Ongoing analyses of mutant forms of these promoter constructs will address the importance of ets binding sites to Lbr gene activation, and binding assays will be used to identify in vivo interactions between GABP and/or PU.1 with the Lbr promoter. We are also generating an EML cell line that contains flox binding sites that flank GABP gene exons, which will be used to generate myeloid progenitors that lack GABP expression. Together our studies are identifying novel mechanisms that regulate the expression of a nuclear envelope protein that is essential for neutrophil development, and may reveal important insight into how morphologic maturation of neutrophils is closely linked to neutrophil functions. Disclosures No relevant conflicts of interest to declare.

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