The yeast two-hybrid system was used to identify binding partners of NuMA, a component of the nuclear matrix in interphase cells. By using the C-terminal half of NuMA as bait, a human cDNA sequence coding for a 223-amino acid protein with a non-helical N-terminal domain and a C-terminal alpha-helical portion was identified and fully sequenced. It was identical to GAS41, a sequence amplified in human gliomas. The sequence of the homologous Drosophila protein was established, and the alignment for GAS41 from nine different species showed that GAS41 is a general eukaryotic protein found in species as diverse as Arabidopsis, Drosophila, Caenorhabditis elegans, yeast, and man. Northern blot analysis showed a single transcript in eight human tissues. A polyclonal antibody to GAS41 showed a dotted staining pattern in interphase nuclei and a uniform distribution in mitotic cells. A GFP-GAS41 fusion protein displayed equivalent patterns. In vitro GAS41 bound to the C-terminal part of the rod region of NuMA, as shown by dot overlay and by surface plasmon resonance measurements. The K(d) of the complex was 2 x 10(-)(7) m. GAS41 is related to the AF-9 and ENL proteins, which are putative transcription factors found as fusion proteins in some acute leukemias. The NuMA/GAS41 interaction may provide a link between nuclear structure and gene expression.

• PMID: 10913114
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Harborth J, Weber K, Osborn M

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