Abstract
Emerin is the gene product of STA whose mutations cause Emery-Dreifuss muscular dystrophy. It is an inner nuclear membrane protein and phosphorylated in a cell cycle-dependent manner. However, the means of phosphorylation of emerin are poorly understood. We investigated the regulation mechanism for the binding of emerin to chromatin, focusing on its cell cycle-dependent phosphorylation in a Xenopus egg cell-free system. It was shown that emerin dissociates from chromatin depending on mitotic phosphorylation of the former, and this plays a critical role in the dissociation of emerin from barrier-to-autointegration factor (BAF). Then, we analyzed the mitotic phosphorylation sites of emerin. Emerin was strongly phosphorylated in an M-phase Xenopus egg cell-free system, and five phosphorylated sites, Ser49, Ser66, Thr67, Ser120, and Ser175, were identified on analysis of chymotryptic and tryptic emerin peptides using a phosphopeptide-concentrating system coupled with a Titansphere column, which specifically binds phosphopeptides, and tandem mass spectrometry sequencing. An in vitro binding assay involving an emerin S175A point mutant protein suggested that phosphorylation at Ser175 regulates the dissociation of emerin from BAF.
Highlights
The nuclear envelope (NE)2 is a highly dynamic structure that disassembles at the onset of mitosis and reassembles on the surface of chromatin during telophase in vertebrates
To determine whether ⌬TM interacts with chromatin and its interaction is regulated in a cell cycle-dependent manner, like for some other inner nuclear membrane proteins, i.e. lamin B receptor (LBR) and LAP2, or not, we performed an in vitro chromatin binding assay
We previously developed this assay method to analyze the binding of inner nuclear membrane proteins to chromatin (28)
Summary
The nuclear envelope (NE) is a highly dynamic structure that disassembles at the onset of mitosis and reassembles on the surface of chromatin during telophase in vertebrates These changes of NE are crucial for cell cycle progression. LAP2 binds to lamin B1 and chromatin, and cell cycle-dependent phosphorylation of LAP2 cancels this binding (4) Phosphorylation of these inner nuclear proteins, is likely to be one of the key mechanisms that control the interactions between the inner nuclear proteins and components of the nuclear lamina as well as chromatin. Emerin belongs to the LEM (LAP2, emerin, MAN1) protein family, whose members have approximately a 40-residue domain named the LEM (10) These proteins directly bind to barrierto-autointegration factor (BAF) (11–13). An in vitro binding assay involving an emerin point mutant revealed that Ser175 phosphorylation is responsible for the dissociation of emerin from BAF
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
