Sun1 forms immobile macromolecular assemblies at the nuclear envelope

Abstract

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.

Structure of Sad1-UNC84 Homology (SUN) Domain Defines Features of Molecular Bridge in Nuclear Envelope

“…Homotrimeric Association of SUN Domain-Previous studies indicate that the SUN1 and SUN2 proteins can undergo homoor heteropolymerization, which involves both the SUN domain and the adjacent coiled coil regions (42,45,49,50). Our current crystallographic study has revealed that the SUN domain alone is sufficient to form a homotrimer (Fig.…”

“…Previous studies showed that SUN1 and SUN2 can form homo-or hetero-oligomers via two predicated coiled coil motifs located in the luminal region near the N-terminal end of the SUN domain (14,30,42,45,49,50). Oligomerization analysis of the potential coiled coil motifs within the luminal domain predicts that although the first coiled coil is highly likely to be dimeric, the second one has a modest probability to be trimeric (58) (supplemental Fig.…”

Structural insights into SUN-KASH complexes across the nuclear envelope

“…These restrictions may facilitate and stabilize higherorder SUN-KASH networks. In addition, SUN proteins could form intermolecular disulfide bonds [15] as well as bind to other yet to be identified proteins, raising the possibility of mechanical tuning across the NE through regulation of the SUN-KASH network. The exact regulatory mechanism of the LINC complex warrants additional investigation in the future.…”

“…Previously, we showed that SUN2 may form homo-oligomers, which is regulated by its two coiled-coil regions and the SUN domain [10]. SUN2 can also form hetero-oligomers with SUN1, another SUN domain protein that is associated with the nuclear pore complex [10,15]. Both SUN1 and SUN2 are widely expressed KASH-binding partners and may have partially redundant functions.…”

Depletion of Nesprin-2 is associated with an embryonic lethal phenotype in mice

“…The largest isoform of Nesprin-2, also known as NUANCE, is an ~ 800 kDa protein (6,885 amino acids) with an N-terminal F-actin binding domain (ABD), a rod domain composed of spectrin repeats and the C-terminal KASH domain[2]. Knockdown studies using shRNAs and overexpression of a shortened protein consisting of the ABD, C-terminal spectrin repeats and the KASH domain revealed that Nesprin-2 is responsible for nuclear size, shape and stability, and determines cell architecture and cell polarization [15–17]. Nesprin-2 is also a component of signaling platforms.…”