emerin and Neuroblastoma

Although several proteins undergo tyrosine phosphorylation at the nuclear envelope, we achieved, for the first time, the identification of tyrosine-phosphorylation sites of a nuclear-membrane protein, emerin, by applying two mass spectrometry-based techniques. With a multiprotease approach combined with highly specific phosphopeptide enrichment and nano liquid chromatography tandem mass spectrometry analysis, we identified three tyrosine-phosphorylation sites, Y-75, Y-95, and Y-106, in mouse emerin. Stable isotope labeling with amino acids in cell culture revealed phosphotyrosines at Y-59, Y-74, Y-86, Y-161, and Y-167 of human emerin. The phosphorylation sites Y-74/Y-75 (human/mouse emerin), Y-85/Y-86, Y-94/Y-95, and Y-105/Y-106 are located in regions previously shown to be critical for interactions of emerin with lamin A, actin or the transcriptional regulators GCL and Btf, while the residues Y-161 and Y-167 are in a region linked to binding lamin-A or actin. Tyrosine Y-94/Y-95 is located adjacent to a five-residue motif in human emerin, whose deletion has been associated with X-linked Emery-Dreifuss muscle dystrophy.

Topics: Amino Acid Sequence; Animals; Cell Culture Techniques; Cell Line; Chromatography, Liquid; HeLa Cells; Humans; Isotope Labeling; Mass Spectrometry; Membrane Proteins; Mice; Molecular Sequence Data; Mutation; Nanotechnology; Neuroblastoma; Nuclear Envelope; Nuclear Proteins; Phosphorylation; Proteomics; Sequence Homology, Amino Acid; Subcellular Fractions; Thymopoietins; Tyrosine