Embryonic and adult isoforms of XLAP2 form microdomains associated with chromatin and the nuclear envelope

Abstract

Laminin-associated polypeptide 2 (LAP2) proteins are alternatively spliced products of a single gene; they belong to the LEM domain family and, in mammals, locate to the nuclear envelope (NE) and nuclear lamina. Isoforms lacking the transmembrane domain also locate to the nucleoplasm. We used new specific antibodies against the N-terminal domain of Xenopus LAP2 to perform immunoprecipitation, identification and localization studies during Xenopus development. By immunoprecipitation and mass spectrometry (LC/MS/MS), we identified the embryonic isoform XLAP2γ, which was downregulated during development similarly to XLAP2ω. Embryonic isoforms XLAP2ω and XLAP2γ were located in close association with chromatin up to the blastula stage. Later in development, both embryonic isoforms and the adult isoform XLAP2β were localized in a similar way at the NE. All isoforms colocalized with lamin B2/B3 during development, whereas XLAP2β was colocalized with lamin B2 and apparently with the F/G repeat nucleoporins throughout the cell cycle in adult tissues and culture cells. XLAP2β was localized in clusters on chromatin, both at the NE and inside the nucleus. Embryonic isoforms were also localized in clusters at the NE of oocytes. Our results suggest that XLAP2 isoforms participate in the maintenance and anchoring of chromatin domains to the NE and in the formation of lamin B microdomains.Electronic supplementary materialThe online version of this article (doi:10.1007/s00441-011-1129-2) contains supplementary material, which is available to authorized users.

Dynamics and Structure-Function Relationships of the Lamin B Receptor (LBR)

“…FL-LBR co-localized to a large extent with nuclear lamin B, but its distribution pattern of was distinct from that of BiP (a luminal protein of the ER) and Nup84/107 (a marker of the NPC) (Fig 1B). In addition, the eGFP fusion protein formed closely spaced microdomains, similar to those identified in previous studies using specific antibodies [16,35]. These raft-like structures, which had contour lengths from 300–600 nm, were readily detectable by both conventional and super-resolution (STED) light microscopy (Fig 1A, detail ).…”

Xenopus LAP2β protein knockdown affects location of lamin B and nucleoporins and has effect on assembly of cell nucleus and cell viability

“…In X. laevis , there have been five cDNA sequences identified that are presumably translated into three different LAP2 proteins discovered so far, ω, β, and γ (Gant et al 1999 , Lang et al 1999 , Lang and Krohne 2003 , Chmielewska et al 2011 ). The Xenopus LAP2β (66 kDa somatic polypeptide) is the only one found in adult animals, whereas two other isoforms, ω and γ (86 and 40 kDa), are present in oocytes and eggs and are downregulated during embryogenesis (Chmielewska et al 2011 ). The N-terminal common fragment of XLAP2, when added to the in vitro nuclear assembly reaction, inhibits chromatin decondensation and nuclear growth similarly to the human LAP2β N-terminal fragment (Gant et al 1999 ).…”

Muscle development, regeneration and laminopathies: how lamins or lamina-associated proteins can contribute to muscle development, regeneration and disease

“…The location of the nucleus is precisely determined within the cell and its disruption may lead to serious dysfunction and decrease in cell viability [48, 140]. This may be exemplified by the phenomenon occurring in syncytial muscle cells in which the majority of nuclei are lined up separately beneath the plasma membrane whereas a small, distinctive population is clustered beneath the neuromuscular junction.…”