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HA95 and LAP2 beta mediate a novel chromatin-nuclear envelope interaction implicated in initiation of DNA replication.

Martins S, Eikvar S, Furukawa K, Collas P - J. Cell Biol. (2003)

Bottom Line: HA95 is a chromatin-associated protein that interfaces the nuclear envelope (NE) and chromatin.Rescue of Cdc6 degradation with proteasome inhibitors restores replication.We propose that an interaction of LAP2beta, or LAP2 proteins, with HA95 is involved in the control of initiation of DNA replication.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medical Biochemistry, University of Oslo, Oslo 0317, Norway.

ABSTRACT
HA95 is a chromatin-associated protein that interfaces the nuclear envelope (NE) and chromatin. We report an interaction between HA95 and the inner nuclear membrane protein lamina-associated polypeptide (LAP) 2 beta, and a role of this association in initiation of DNA replication. Precipitation of GST-LAP2 beta fusion proteins and overlays of immobilized HA95 indicate that a first HA95-binding region lies within amino acids 137-242 of LAP2 beta. A second domain sufficient to bind HA95 colocalizes with the lamin B-binding domain of LAP2beta at residues 299-373. HA95-LAP2 beta interaction is not required for NE formation. However, disruption of the association of HA95 with the NH2-terminal HA95-binding domain of LAP2 beta abolishes the initiation, but not elongation, of DNA replication in purified G1 phase nuclei incubated in S-phase extract. Inhibition of replication initiation correlates with proteasome-mediated proteolysis of Cdc6, a component of the prereplication complex. Rescue of Cdc6 degradation with proteasome inhibitors restores replication. We propose that an interaction of LAP2beta, or LAP2 proteins, with HA95 is involved in the control of initiation of DNA replication.

Show MeSH
Functional domains of LAP2β. Identified functional regions of LAP2β (LEM and transmembrane [TM]) and domains binding to indicated ligands are shown. Numbers refer to amino acid positions. See text for references to binding domains.
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fig9: Functional domains of LAP2β. Identified functional regions of LAP2β (LEM and transmembrane [TM]) and domains binding to indicated ligands are shown. Numbers refer to amino acid positions. See text for references to binding domains.

Mentions: This study provides evidence for a novel direct interaction of the NE with the genome, via the INM protein LAP2β and the chromatin- and nuclear matrix–associated protein HA95. The nucleoplasmic 410 amino acids of LAP2β are prompt to interactions with multiple intranuclear ligands (Fig. 9). The first 50 residues are common to all LAP2 proteins and bind DNA (Cai et al., 2001), in consistency with the chromatin-binding property of this region, which causes post-mitotic association of LAP2α with chromosomes (Vlcek et al., 1999). LAP2 proteins also bind the DNA-bridging protein BAF through residues 67–137 (Furukawa, 1999; Shumaker et al., 2001), which include most of the LEM domain (Fig. 9). The significance of this interaction in interphase remains unclear, although a role in chromatin decondensation and nuclear expansion in vitro has been proposed (Segura-Totten et al., 2002). LAP2β also binds GCL, a transcription regulator that interacts with components of the E2F transcription machinery (Nili et al., 2001). LAP2β is capable of reducing the transcription activity of the E2F complex alone or with GCL (Nili et al., 2001), suggesting an involvement of the NE in transcription regulation. It will be interesting to determine whether mutations in NE proteins that cause disease (Vigouroux and Bonne, 2002) also lead to defects in transcription regulation in affected tissues.


HA95 and LAP2 beta mediate a novel chromatin-nuclear envelope interaction implicated in initiation of DNA replication.

Martins S, Eikvar S, Furukawa K, Collas P - J. Cell Biol. (2003)

Functional domains of LAP2β. Identified functional regions of LAP2β (LEM and transmembrane [TM]) and domains binding to indicated ligands are shown. Numbers refer to amino acid positions. See text for references to binding domains.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172640&req=5

fig9: Functional domains of LAP2β. Identified functional regions of LAP2β (LEM and transmembrane [TM]) and domains binding to indicated ligands are shown. Numbers refer to amino acid positions. See text for references to binding domains.
Mentions: This study provides evidence for a novel direct interaction of the NE with the genome, via the INM protein LAP2β and the chromatin- and nuclear matrix–associated protein HA95. The nucleoplasmic 410 amino acids of LAP2β are prompt to interactions with multiple intranuclear ligands (Fig. 9). The first 50 residues are common to all LAP2 proteins and bind DNA (Cai et al., 2001), in consistency with the chromatin-binding property of this region, which causes post-mitotic association of LAP2α with chromosomes (Vlcek et al., 1999). LAP2 proteins also bind the DNA-bridging protein BAF through residues 67–137 (Furukawa, 1999; Shumaker et al., 2001), which include most of the LEM domain (Fig. 9). The significance of this interaction in interphase remains unclear, although a role in chromatin decondensation and nuclear expansion in vitro has been proposed (Segura-Totten et al., 2002). LAP2β also binds GCL, a transcription regulator that interacts with components of the E2F transcription machinery (Nili et al., 2001). LAP2β is capable of reducing the transcription activity of the E2F complex alone or with GCL (Nili et al., 2001), suggesting an involvement of the NE in transcription regulation. It will be interesting to determine whether mutations in NE proteins that cause disease (Vigouroux and Bonne, 2002) also lead to defects in transcription regulation in affected tissues.

Bottom Line: HA95 is a chromatin-associated protein that interfaces the nuclear envelope (NE) and chromatin.Rescue of Cdc6 degradation with proteasome inhibitors restores replication.We propose that an interaction of LAP2beta, or LAP2 proteins, with HA95 is involved in the control of initiation of DNA replication.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medical Biochemistry, University of Oslo, Oslo 0317, Norway.

ABSTRACT
HA95 is a chromatin-associated protein that interfaces the nuclear envelope (NE) and chromatin. We report an interaction between HA95 and the inner nuclear membrane protein lamina-associated polypeptide (LAP) 2 beta, and a role of this association in initiation of DNA replication. Precipitation of GST-LAP2 beta fusion proteins and overlays of immobilized HA95 indicate that a first HA95-binding region lies within amino acids 137-242 of LAP2 beta. A second domain sufficient to bind HA95 colocalizes with the lamin B-binding domain of LAP2beta at residues 299-373. HA95-LAP2 beta interaction is not required for NE formation. However, disruption of the association of HA95 with the NH2-terminal HA95-binding domain of LAP2 beta abolishes the initiation, but not elongation, of DNA replication in purified G1 phase nuclei incubated in S-phase extract. Inhibition of replication initiation correlates with proteasome-mediated proteolysis of Cdc6, a component of the prereplication complex. Rescue of Cdc6 degradation with proteasome inhibitors restores replication. We propose that an interaction of LAP2beta, or LAP2 proteins, with HA95 is involved in the control of initiation of DNA replication.

Show MeSH