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Henning Otto

Publications and source records attributed to Henning Otto.

5 recordsLinked to original sources
PubMed2006

Identification of tyrosine-phosphorylation sites in the nuclear membrane protein emerin.

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.

Amino Acid Sequence↗
PubMed2004

Sun2 is a novel mammalian inner nuclear membrane protein.

Sun protein (Sun1 and Sun2) cDNAs were previously cloned based on the homology of their C-terminal regions (SUN (Sad1 and UNC) domain) with the Caenorhabditis elegans protein UNC-84 whose mutation disrupts nuclear migration/positioning. In this study, we raised an anti-Sun2 serum and identified Sun2 in mammalian cells. In HeLa cells, Sun2 displays a nuclear rim-like pattern typical for a nuclear envelope protein. The Sun2 antibody signal co-localizes with nuclear pore and INM markers signals. The rim-like pattern was also observed with the recombinant full-length Sun2 protein fused to either EGFP or V5 epitopes. In addition, we found that a recombinant truncated form of Sun2, extending from amino acids 26 to 339, is sufficient to specify the nuclear envelope localization. Biochemical analyses show that Sun2 is an 85-kDa protein that is partially insoluble in detergent with high salt concentration and in chaotropic agents. Furthermore, Sun2 is enriched in purified HeLa cell nuclei. Electron microscopy analysis shows that Sun2 localizes in the nuclear envelope with a sub-population present in small clusters. Additionally, we show that the SUN domain of Sun2 is localized to the periplasmic space between the inner and the outer nuclear membranes. From our data, we conclude that Sun2 is a new mammalian inner nuclear membrane protein. Because the SUN domain is conserved from fission yeast to mammals, we suggest that Sun2 belongs to a new class of nuclear envelope proteins with potential relevance to nuclear membrane function in the context of the involvement of its components in an increasing spectrum of human diseases.

Animals↗
PubMed2004

A surface-modified dendrimer set for potential application as drug delivery vehicles: synthesis, in vitro toxicity, and intracellular localization.

The synthesis, cytotoxicity, and behavior in cell culture of a new set of first- (G1) and second-generation (G2) dendrimers is reported. The surface functionality of these dendrimers has been varied to see whether structure/toxicity relations can be observed. The outermost functional groups are amines that are decorated either with protons, tert-butoxycarbonyl (Boc) or benzyloxycarbonyl (Cbz) protecting groups, Boc-protected or unprotected natural amino acid residues, ethylenediamine ligands, and/or dansyl fluorescence labels. The cytotoxicity was determined in vitro in concentration-dependent assays using the human MCF-7 breast cancer cell line. Cellular uptake and intracellular distribution was monitored by confocal fluorescence microscopy after internalization of the dansyl-labeled dendrimers by HeLa cells.

Amino Acids↗