A lamin B receptor in the nuclear envelope.

Abstract

Using a solution binding assay, we show that purified 1251-labeled lamin B binds in a saturable and specific fashion to lamin-depleted avian erythrocyte nuclear membranes with a Kd of -0.2 ,LM. This binding is significantly greater than the binding of 12'I-labeled lamin A and is competitively inhibited by unlabeled ligand. We demonstrate that a 58-kDa integral membrane protein (p58) is a lamin B receptor by virtue of its abundance in the nuclear envelope and association with '251-labeled lamin B in ligand blotting assays. Specific antibodies raised against p58 recognize one protein in isolated nuclei and partially block 1251-labeled lamin B binding to lamin-depleted nuclear membranes. Cell fractionation and indirect immunofluorescence microscopy show that p58 is located in the periphery of the nucleus. This protein may serve as a membrane attachment site for the nuclear lamina by acting as a specific receptor for lamin B.The nuclear lamina is a filamentous meshwork situated between the inner nuclear membrane and heterochromatin (1-3). This structure provides support for the adjacent inner nuclear membrane (4) and possible attachment sites for chromatin (2, 5, 6) and cytoplasmic intermediate filaments (7). The nuclear lamins, a family of interrelated polypeptides structurally homologous to intermediate filament proteins, are the building blocks of the nuclear lamina (3,(8)(9)(10). Two major types of lamins have been recognized in higher eukaryotic cells: the acidic B-lamins and the neutral A-lamins (10-12). During mitosis and lamina depolymerization, the B-lamins remain associated with intracellular membrane vesicles thought to be remnants of the inner nuclear membrane, whereas the A-lamins become soluble (12). The B-lamins are also more resistant to extraction from the nuclear envelope compared to the A-lamins (13). Because of these findings, the B-lamins are thought to mediate the coupling of the lamina to the inner nuclear membrane (5, 13). It has been proposed that lamin B associates with the membrane either by a hydrophobic interaction with components of the lipid bilayer (14) or by an attachment to a transmembrane receptor protein (2, 13). These questions and controversies prompted the present investigations. MATERIALS AND METHODSCell Fractionation and Protein Purification. Nuclear envelopes and plasma membranes from turkey erythrocytes were prepared as previously described (7). Lamin-depleted envelopes were prepared by extraction with 8 M urea/10 mM Tris HCl, pH 8.0/1 mM EDTA (see below). Lamins A and B were purified from the urea extract of turkey or rat nuclear envelopes by DEAE-cellulose chromatography and were '25I-labeled with Bolton-Hunter reagent as described (7).Extractions with (i) 8 M urea/10 mM Tris HCl, pH 8.0/1 mM EDTA, (ii) 0.10 M Na2CO3 (pH 11.5), and (iii) 2% Triton X-100/2 M KCl/10 mM Tris-HCl, pH 8.0, were performed on nuclear envelopes at room temperature. All extraction solutions also contained 1 mM dithiothreitol and 0.2 mM phenylmethylsulfonyl fluoride. Bath sonication was use...

A study of goat SRY protein expression suggests putative new roles for this gene in the developing testis of a species with long‐lasting SRY expression

“…The peri-nuclear localization appeared to be very similar to that observed with lamin B (Fig. 4D,F), a receptor located in the nuclear membrane (Worman et al, 1988), but unlike lamin B, cells outside the gonad did not display any peri-nuclear SRY staining (compare Fig. 4C with 4F).…”

The inner nuclear membrane protein Lem2 is critical for normal nuclear envelope morphology

“…To obtain insight into the function of INM proteins in NE integrity we performed RNAi on the transmembrane proteins lamin B receptor (LBR) [16], lamina-associated polypeptide 2b (Lap2b) [4], emerin [17] and Lem2. Only depletion of Lem2 led to a detectable phenotype (see below).…”

Inner nuclear membrane and signal transduction

“…Binding to fixed nuclear structures is responsible for the retention of proteins in the inner nuclear membrane Worman, 1993, 1995;Ellenberg et al, 1997;Ö stlund et al, 1999;Ohba et al, 2004]. Most of the characterized integral proteins of the inner nuclear membrane bind to nuclear lamins and/or chromatin components, example being lamin B receptor [Worman et al, 1988;Worman, 1994, 1996;Ye et al, 1997] and isoforms of lamina-associated polypeptide 2 [Foisner and Gerace, 1993]. Data obtained from a proteomics analysis of purified nuclear envelopes suggest that approximately 80 transmembrane proteins reside in the inner nuclear membrane of interphase cells [Schirmer et al, 2003].…”