Nuclear positioning is central to many cellular and developmental events. A bridge of inner nuclear membrane (INM) SUN proteins and outer nuclear membrane (ONM) KASH proteins spans the nuclear envelope. In C. elegans, the SUN protein UNC-84 recruits the KASH protein UNC-83 to the ONM where it targets dynein and kinesin-1 to move nuclei. However, it remains unknown how SUN proteins span the perinuclear space of the nuclear envelope and how forces transferred across the bridge are dissipated at the nucleoskeleton. To elucidate mechanisms of how mechanical forces are transferred from the bridge to the nucleoskeleton, we performed a yeast two-hybrid screen with the nucleoplasmic domain of UNC-84 as bait and identified an interaction with the lamin B gene LMN-1.
lmn-1(RNAi) animals had a
hyp7 nuclear migration defect. Moreover, the LMN-1 interaction was significantly weakened by an UNC-84 P91S mutation that was previously identified in genetic screens with a partial loss-of-function
hyp7 nuclear migration defect. These data support a model where UNC-84 interacts with LMN-1 to dissipate force from the bridge to the nucleoskeleton. We identified a new player in this network, the integral INM protein SAMP-1 (T24F1.2);
samp-1(RNAi) animals had a weak nuclear migration defect. To test the hypothesis that UNC-84 spans and regulates the space between the INM and ONM, we examined the morphology of the nuclear envelope by electron microscopy in
unc-84(null) mutants. We observed extreme separation of the ONM from the INM in
unc-84(null) L1 muscle nuclei. Surprisingly, a deletion of ~300 residues between the trans-membrane and SUN domains of UNC-84 remained functional, thus, the majority of this linker domain is not required for nuclear migration. However, a 60 amino acid span immediately upstream of the SUN domain, and the SUN domain itself are required for function. Additionally, mutations in the SUN domain of UNC-84 predicted to interact with KASH peptides from human SUN structures disrupt
hyp7 nuclear migrations. In our model the nucleoplasmic domain of UNC-84 directly binds lamin to transfer forces from the bridge to the nucleoskeleton. UNC-84 is also required for even nuclear envelope morphology.