Characterization of the topology, targeting and binding properties of SUN proteins and their involvement in laminopathies
thesisposted on 18.01.2011, 15:31 by Farhana Haque
The nuclear envelope (NE) is a double membrane structure enclosing the chromatin and forms the interface between cytoplasm and nucleus. The NE harbours numerous integral membrane proteins, mostly located at the inner nuclear membrane (INM) and is underlined by the nuclear lamina, which along with its interacting proteins gives mechanical strength to the NE. Mutation in lamins and their interacting NE proteins give rise to laminopathy diseases. SUN1 and SUN2 are novel mammalian NE proteins, sharing a conserved C-terminal SUN domain. Their C. elegans homologue, UNC-84, is hypothesized to have roles in nuclear migration and positioning by forming a bridge across the NE through interaction with ANC-1 in the outer nuclear membrane (ONM), which in turn binds cytoplasmic actin. Previously, SUN1 was identified in the lab as a lamin A-binding protein in a yeast two-hybrid screen. Here, I have confirmed SUN1 as an INM protein, comprising a nucleoplasmic N-terminus that interacts with lamin A and a lumenal C-terminus that interacts with mammalian ANC-1 homologues, nesprins. I further identified novel nucleoplasmic interactions of the SUN proteins with emerin and nucleoplasmic isoforms of nesprins, thus demonstrating multi-protein interactions of SUN proteins at the NE. Notably, lamin A/C, emerin and nesprins are mutated in the laminopathy Emery-Dreifuss muscular dystrophy (EDMD). Here, I have demonstrated SUN protein involvement with laminopathies for the first time. SUN1 and SUN2 interaction with L530P and L527P EDMD lamin A mutants and with G608G and T623S progeria lamin A mutants were dramatically reduced. Although SUN proteins were not mislocalized from the NE in EDMD patient fibroblasts examined, increased recruitment of SUN1, but not SUN2, was observed in progeria patient fibroblasts, possibly due to increased expression of prelamin A. Subtle disruptions in the interactions between SUN proteins and their binding partners may therefore contribute to laminopathy disease phenotypes.