LBR, a chromatin and lamin binding protein from the inner nuclear membrane, is proteolyzed at late stages of apoptosis

1998 ◽  
Vol 111 (10) ◽  
pp. 1441-1451 ◽  
Author(s):  
I. Duband-Goulet ◽  
J.C. Courvalin ◽  
B. Buendia
Keyword(s):  
Nuclear Membrane ◽  
Binding Protein ◽  
Transmembrane Protein ◽  
Chromatin Condensation ◽  
Lamin B ◽  
Inner Nuclear Membrane ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Amino Terminal Domain ◽  
Late Stages

Chromatin condensation and apposition to the nuclear envelope is an important feature of the execution phase of apoptosis. During this process, lamin proteins that are located between the inner nuclear membrane and heterochromatin are proteolyzed by the apoptosis-specific protease caspase 6. We have investigated the fate of nuclear membranes during apoptosis by studying the lamin B receptor (LBR), a transmembrane protein of the inner nuclear membrane. LBR interacts through its nucleoplasmic amino-terminal domain with both heterochromatin and B-type lamins, and is phosphorylated throughout the cell cycle, but on different sites in interphase and mitosis. We report here that: (i) the amino-terminal domain of LBR is specifically cleaved during apoptosis to generate an approximately 20 kDa soluble fragment; (ii) the cleavage of LBR is a late event of apoptosis and occurs subsequent to lamin B cleavage; (iii) the phosphorylation of LBR during apoptosis is similar to that occurring in interphase. As the association of condensed chromatin with the inner nuclear membrane persists until the late stages of apoptosis, we suggest that the chromatin binding protein LBR plays a major role in maintaining this association.

scholarly journals Signals and structural features involved in integral membrane protein targeting to the inner nuclear membrane.

1995 ◽  
Vol 130 (1) ◽  
pp. 15-27 ◽  
Author(s):  
B Soullam ◽  
H J Worman
Keyword(s):  
Membrane Protein ◽  
Nuclear Membrane ◽  
Structural Features ◽  
Inner Nuclear Membrane ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Integral Protein ◽  
Carboxyl Terminal Domain ◽  
Carboxyl Terminal ◽  
Amino Terminal Domain

We have examined transfected cells by immunofluorescence microscopy to determine the signals and structural features required for the targeting of integral membrane proteins to the inner nuclear membrane. Lamin B receptor (LBR) is a resident protein of the nuclear envelope inner membrane that has a nucleoplasmic, amino-terminal domain and a carboxyl-terminal domain with eight putative transmembrane segments. The amino-terminal domain of LBR can target both a cytosolic protein to the nucleus and a type II integral protein to the inner nuclear membrane. Neither a nuclear localization signal (NLS) of a soluble protein, nor full-length histone H1, can target an integral protein to the inner nuclear membrane although they can target cytosolic proteins to the nucleus. The addition of an NLS to a protein normally located in the inner nuclear membrane, however, does not inhibit its targeting. When the amino-terminal domain of LBR is increased in size from approximately 22.5 to approximately 70 kD, the chimeric protein cannot reach the inner nuclear membrane. The carboxyl-terminal domain of LBR, separated from the amino-terminal domain, also concentrates in the inner nuclear membrane, demonstrating two nonoverlapping targeting signals in this protein. Signals and structural features required for the inner nuclear membrane targeting of proteins are distinct from those involved in targeting soluble polypeptides to the nucleoplasm. The structure of the nucleocytoplasmic domain of an inner nuclear membrane protein also influences targeting, possibly because of size constraints dictated by the lateral channels of the nuclear pore complexes.

Interaction between the inner nuclear membrane lamin B receptor and the heterochromatic methyl binding protein, MeCP2

2009 ◽  
Vol 315 (11) ◽  
pp. 1895-1903 ◽  
Author(s):  
Alessia Guarda ◽  
Fabrizio Bolognese ◽  
Ian Marc Bonapace ◽  
Gianfranco Badaracco
Keyword(s):  
Nuclear Membrane ◽  
Binding Protein ◽  
Lamin B ◽  
Inner Nuclear Membrane ◽  
Lamin B Receptor ◽  
Protein Mecp2

scholarly journals Self-association of the Amino-terminal Domain of the Yeast TATA-binding Protein

2003 ◽  
Vol 279 (2) ◽  
pp. 1376-1382 ◽  
Author(s):  
Claire A. Adams ◽  
Sambit R. Kar ◽  
James E. Hopper ◽  
Michael G. Fried
Keyword(s):  
Binding Protein ◽  
Tata Binding Protein ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Self Association ◽  
Amino Terminal Domain

Subcellular distribution of the Xenopus p58/lamin B receptor in oocytes and eggs

1999 ◽  
Vol 112 (15) ◽  
pp. 2583-2596 ◽  
Author(s):  
A. Gajewski ◽  
G. Krohne
Keyword(s):  
Nuclear Membrane ◽  
Xenopus Oocytes ◽  
Immunofluorescence Microscopy ◽  
Significant Proportion ◽  
Sperm Chromatin ◽  
Lamin B ◽  
Inner Nuclear Membrane ◽  
Amino Terminal ◽  
Lamin B Receptor

The p58/lamin B receptor of vertebrates is localized in the inner nuclear membrane. Antibodies raised against the bacterially expressed amino-terminal half of Xenopus p58 (Xp58) revealed that in Xenopus oocytes the vast majority of this membrane protein is localized in cytoplasmic membranes. Only very small amounts of p58 not detectable by immunofluorescence microscopy were contained in the oocyte nuclear envelope. In contrast, nuclear membranes of 2-cell stage embryos were successfully stained with p58 antibodies, nuclei reconstituted in vitro in Xenopus egg extracts contained p58, and the nucleoplasmic domain of Xp58 could be specifically bound to sperm chromatin in vitro. One major difference between oocytes and early embryonic cells is that no chromatin is associated with the oocyte inner nuclear membrane whereas the complement of lamins is identical in both cell types. To gain insight into the properties of oocyte p58 we microinjected isolated nuclei of cultured rat cells into the cytoplasm of Xenopus oocytes. The oocyte p58 was detectable by immunofluorescence microscopy within 16–20 hours in the nuclear membrane of rat nuclei. Our data indicate that the peripheral chromatin but not lamins are required for the retention of p58 in the inner nuclear membrane. Sucrose step gradient centrifugation of total oocyte membranes revealed that the oocyte p58 was predominantly recovered in membrane fractions that did not contain lamins whereas membrane associated lamins and p58 of unfertilized eggs were found in the same fractions. By electron microscopical immunolocalizations one major population of meiotic p58 vesicles was identified that contained exclusively p58 and a second minor population (ca. 11% of p58 vesicles) contained in addition to p58 membrane bound B-type lamins. Egg vesicles containing pore membrane proteins were predominantly recovered in gradient fractions that did not contain p58 and B-type lamins. Our data indicate that the targeting of p58 to chromatin at the end of mitosis in the early Xenopus embryo is a process independent from that of lamin targeting. Comparable to the situation in oocytes and eggs, a significant proportion of p58 of interphase cells could be recovered in fractions that did not contain lamins. This population of p58 molecules could be extracted from A6-cells with buffers containing 1% Triton X-100/0.15 M NaCl and could be pelleted by a 50,000 g centrifugation. A- and B-type lamins were not detectable in the p58 containing pellet.

scholarly journals Tethering of an E3 ligase by PCM1 regulates the abundance of centrosomal KIAA0586/Talpid3 and promotes ciliogenesis

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Lei Wang ◽  
Kwanwoo Lee ◽  
Ryan Malonis ◽  
Irma Sanchez ◽  
Brian D Dynlacht
Keyword(s):  
Binding Protein ◽  
E3 Ligase ◽  
Human Cells ◽  
Gene Encoding ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Integral Component ◽  
Specific Proteins ◽  
Amino Terminal Domain

To elucidate the role of centriolar satellites in ciliogenesis, we deleted the gene encoding the PCM1 protein, an integral component of satellites. PCM1 null human cells show marked defects in ciliogenesis, precipitated by the loss of specific proteins from satellites and their relocation to centrioles. We find that an amino-terminal domain of PCM1 can restore ciliogenesis and satellite localization of certain proteins, but not others, pinpointing unique roles for PCM1 and a group of satellite proteins in cilium assembly. Remarkably, we find that PCM1 is essential for tethering the E3 ligase, Mindbomb1 (Mib1), to satellites. In the absence of PCM1, Mib1 destabilizes Talpid3 through poly-ubiquitylation and suppresses cilium assembly. Loss of PCM1 blocks ciliogenesis by abrogating recruitment of ciliary vesicles associated with the Talpid3-binding protein, Rab8, which can be reversed by inactivating Mib1. Thus, PCM1 promotes ciliogenesis by tethering a key E3 ligase to satellites and restricting it from centrioles.

scholarly journals The amino-terminal domain of the lamin B receptor is a nuclear envelope targeting signal.

1993 ◽  
Vol 120 (5) ◽  
pp. 1093-1100 ◽  
Author(s):  
B Soullam ◽  
H J Worman
Keyword(s):  
Nuclear Envelope ◽  
Chimeric Protein ◽  
Integral Membrane Protein ◽  
Lamin B ◽  
Amino Terminal ◽  
Transmembrane Segments ◽  
Terminal Domain ◽  
Lamin B Receptor ◽  
Targeting Signal ◽  
Amino Terminal Domain

The lamin B receptor (LBR) is a polytopic protein of the inner nuclear membrane. It is synthesized without a cleavable amino-terminal signal sequence and composed of a nucleoplasmic amino-terminal domain of 204 amino acids followed by a hydrophobic domain with eight putative transmembrane segments. To identify a nuclear envelope targeting signal, we have examined the cellular localization by immunofluorescence microscopy of chicken LBR, its amino-terminal domain and chimeric proteins transiently expressed in transfected COS-7. Full-length LBR was targeted to the nuclear envelope. The amino-terminal domain, without any transmembrane segments, was transported to the nucleus but excluded from the nucleolus. When the amino-terminal domain of LBR was fused to the amino-terminal side of a transmembrane segment of a type II integral membrane protein of the ER/plasma membrane, the chimeric protein was targeted to the nuclear envelope, likely the inner nuclear membrane. When the amino-terminal domain was deleted from LBR and replaced by alpha-globin, the chimeric protein was retained in the ER. These findings demonstrate that the amino-terminal domain of LBR is targeted to the nucleus after synthesis in the cytoplasm and that this polypeptide can function as a nuclear envelope targeting signal when located at the amino terminus of a type II integral membrane protein synthesized on the ER.

scholarly journals The first membrane spanning region of the lamin B receptor is sufficient for sorting to the inner nuclear membrane.

1993 ◽  
Vol 120 (3) ◽  
pp. 631-637 ◽  
Author(s):  
S Smith ◽  
G Blobel
Keyword(s):  
Nuclear Membrane ◽  
Mammalian Cells ◽  
Specific Interaction ◽  
Integral Membrane Protein ◽  
Transmembrane Segment ◽  
Hydrophobic Domain ◽  
Lamin B ◽  
Inner Nuclear Membrane ◽  
Amino Terminal ◽  
Lamin B Receptor

The lamin B receptor (LBR) is a polytopic integral membrane protein localized exclusively in the inner nuclear membrane domain of the nuclear envelope. Its cDNA deduced primary structure consists of a highly charged amino-terminal domain of 205 residues that faces the nucleoplasm followed by a hydrophobic domain with eight potential transmembrane segments. To identify determinants that sort LBR from its site of integration (RER and outer nuclear membrane) to the inner nuclear membrane, we prepared full-length, truncated, and chimeric cDNA constructs of chick LBR, transfected these into mammalian cells and detected the expressed protein by immunofluorescence microscopy using appropriate antibodies. Surprisingly, we found that the determinants for sorting of LBR to the inner nuclear membrane reside in a region comprising its first transmembrane sequence plus flanking residues on either side. The other transmembrane regions as well as the nucleoplasmic domain are not required for sorting. We propose that the first transmembrane segment of LBR interacts specifically with another transmembrane segment and consider several mechanisms by which such specific interaction could result in sorting to the inner nuclear membrane.

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