Sun1 forms immobile macromolecular assemblies at the nuclear envelope.
SUN-domain proteins form a novel and conserved family of inner nuclear membrane (INM) proteins, which establish physical connections between the nucleoplasm and the cytoskeleton. In the current study, we provide evidence that within the nuclear envelope (NE) Sun1 proteins form highly immobile oligomeric complexes in interphase cells. By performing inverse fluorescence recovery after photobleaching analysis, we demonstrate in vivo that both perinuclear and nucleoplasmic Sun1 segments are essential for maintenance of Sun1 immobility at the NE. Our data in particular underline the self-association properties of the C-terminal coiled-coil Sun1 segment, the ability of which to form dimers and tetramers is demonstrated. Furthermore, the Sun1 tertiary structure involves interchain disulfide bonds that might contribute to higher homo-oligomer formation, although the overall dynamics of the Sun1 C-terminus remains unaffected when the cysteins involved are mutated. While a major Sun1 pool colocalizes with nuclear pore complex proteins, a large fraction of the Sun1 protein assemblies colocalize with immunoreactive foci of Sun2, another SUN-domain paralogue at the NE. We demonstrate that the Sun1 coiled-coil domain permits these heterophilic associations with Sun2. Sun1 therefore provides a non-dynamic platform for the formation of different macromolecular assemblies at the INM. Our data support a model in which SUN-protein-containing multi-variate complexes may provide versatile outer nuclear membrane attachment sites for cytoskeletal filaments.
Center for Biochemistry, Medical Faculty, University of Cologne, Joseph-Stelzmann-Strasse 52, 50931 Cologne, Germany.
Wenshu
W
Lu
Josef
J
Gotzmann
Lucia
L
Sironi
Verena-Maren
VM
Jaeger
Maria
M
Schneider
Yvonne
Y
Lüke
Mathias
M
Uhlén
Cristina Al-Khalili
CA
Szigyarto
Andreas
A
Brachner
Jan
J
Ellenberg
Roland
R
Foisner
Angelika A
AA
Noegel
Iakowos
I
Karakesisoglou
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002467:Cell Nucleus / Q000378:metabolism* / Q000648:ultrastructure; D003432:Cross-Linking Reagents; D004220:Disulfides / Q000378:metabolism; D036681:Fluorescence Recovery After Photobleaching; D005455:Fluorescent Antibody Technique; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007124:Immunoenzyme Techniques; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009685:Nuclear Envelope / Q000378:metabolism* / Q000648:ultrastructure; D022022:Nuclear Pore / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques
D003432:Cross-Linking Reagents; D004220:Disulfides; D008869:Microtubule-Associated Proteins; D011993:Recombinant Fusion Proteins; C464932:SUN1 protein, mouse; D005982:Glutathione Transferase